bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B40: Methylation signatures specific to triple negative breast cancer subtypes
Authors: Kowalski Jeanne , Switchenko Jeffery M. , Dwivedi Bhakti , Nickleach Dana , Moreno Carlos S.
Published: August 2014

Objective: We identify methylation signatures specific to triple negative breast cancer (TNBC) subtypes using publicly available data, and examine their association with expression.

Materials and Methods: DNA methylation, RNAseq expression and clinical data from breast invasive carcinoma patients were obtained from The Cancer Genome Atlas (TCGA) Data portal. The classification of each patient sample as TNBC was defined according to a combination of FISH and/or IHC results for ER, PR and HER2 negativity. Each TNBC specimen was classified into one of the following molecular subtypes by applying the gene expression-based model of Lehman (2011): basal-like 1 (BL1), basal-like 2 (BL2), immunomodulatory (IM), mesenchymal (MES), mesenchymal stem-like (MSL), luminal androgen receptor (LAR), and unstable (UNS). Among the 116 identified TNBC specimens, 61 had level-three data available on both methylation and RNAseq expression. For methylation, beta values were extracted based on Illumina's Infinium Human DNA Methylation 450K platform; for expression, RNAseqV2 Expectation/Maximization normalized counts (RSEM: Li and Dewey, 2011) were extracted on 20,531 genes and a logarithmic transformation applied. Data were filtered to identify methylation changes specific to each TNBC subtype based on a novel, generalized statistic (Switchenko and Kowalski, 2013) and tested for median difference between subtypes using p < 0.10 to define significance.

Results: Among the 61 TNBC samples, the following molecular subtypes were represented: BL1 (n=6), BL2 (n=6), IM (n=12), LAR (n=5), MES (n=15), MSL (n=4), and UNS (n=13). A number of genes were determined to be hypermethylated with corresponding decreased expression by RNAseq in several subtypes. Methylated genes specific to the LAR TNBC subtype included IGF1R (61% LAR vs. 2% other) and N4BP2L1 (44% LAR vs. 3% other). Methylated genes in the MES subtype included ACSL5 (80% vs. 50%), RHOH (82% vs. 6%) and OAS2 (26% vs. 7%), and specific to the BL2 subtype were EN1 (35% vs. 8%) and TRAFIP3 (60% vs. 35%). Two hypomethylated genes, PIK3CD and CD69, associated with respective subtypes, MSL and IM, showed corresponding increased expression levels.

Conclusion: Our results provide further insight into the molecular heterogeneity surrounding TNBC patients by examining methylation changes specific to molecular subtypes, which may be further examined for their effect on clinical outcome.

Citation Format: Jeanne Kowalski, Jeffery M. Switchenko, Bhakti Dwivedi, Dana Nickleach, Carlos S. Moreno. Methylation signatures specific to triple negative breast cancer subtypes. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr B40.
Abstract B64: Conglomerate methylation silencing of clustered proto-cadherin genes in the course of cervical carcinogenesis
Authors: Chu Tang-Yuan , Wang Kai-Hung , Lin Cuei-Jyuan
Published: August 2014

Objective: A common phenomenon in cancer invasion involves global epigenetic remodeling of cell adhesion genes. Taking the advantage of the discrete spectrum of lesions in cervical carcinogenesis, this study aims to discover and characterize globally methylated cell adhesion genes in the early invasion period of cervical carcinogenesis.

Methods: Genome-wide methylation array was performed on 36 cervical intraepithelial neoplasia (CIN) 3/ early invasive cancer (CA) and 36 normal/inflammation/CIN1 specimens. Methylation silencing was validated in cancer cells and cohort specimens by bisulfite sequencing, methylation specific PCR, RT-PCR and 5-Aza-dC treatment.

Results: Widespread methylation silencing of 12 members of clustered protocadherin (PCDH) genes including PCDHA4, PCDHA8, PCDHA10, PCDHA13, PCDHB3, PCDHB6, PCDHB7, PCDHB14, PCDH GA12, PCDHGB6, PCDHGB7 and PCDHGC3 was discovered and validated. These PCDH genes were collectively silenced by methylation in differing spectra in a wide variety of cancer cells including carcinoma of the cervix (4/4 cell lines), endometrium (2/2), liver (3/3), head and neck (3/3), breast (1/3), and lung (1/2). In an independent cohort of the spectrum of cervical neoplasia including 60 normals, 30 CIN1, 20 CIN2/3 and 80 CA, methylated PCDHA13 was noted in 17%, 58%, 69% and 78%, respectively (P<10-11).

Conclusions: Methylation silencing of clustered PCDH genes is a general phenomenon of carcinomas. Methylation of members of this gene family can be valuable diagnostic markers in detection of cancers as exemplified by methylated PCDHA13 in cervical cancer.

Citation Format: Tang-Yuan Chu, Kai-Hung Wang, Cuei-Jyuan Lin. Conglomerate methylation silencing of clustered proto-cadherin genes in the course of cervical carcinogenesis. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr B64.
Abstract IA01: Transcriptional and epigenetic control of oncogenes
Authors: Young Richard A.
Published: August 2014

The gene expression program of any particular cell largely defines its identity and its functions. I will describe new insights into the roles that transcription factors and chromatin regulators play in the control of gene expression programs during normal development and in cancer. These insights provide opportunities for the development of novel therapeutics for cancer and other diseases.

Citation Format: Richard A. Young. Transcriptional and epigenetic control of oncogenes. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA01.
Abstract IA07: New functions for histone modifying enzymes
Authors: Dent Sharon Y.R. , Atanassov Boyko , Hirsch Calley , Koutelou Evangelia , Latham John
Published: August 2014

Histone modifying enzymes are important creating and maintaining epigenetic programs that regulate cell identity and growth. Mutations or abnormal expression of these proteins are associated with multiple cancers. Our lab uses genetic approaches to define the full spectrum of functions for these enzymes. For example, we have created a series of mutations in the mouse Gcn5 gene in order to define the functions of this histone acetyltransferase (HAT) in a mammalian system. Deletion of Gcn5 led to early embryonic death and to telomere dysfunction. Biochemical studies revealed that depletion of Gcn5 or ubiquitin specific protease 22 (Usp22), which is another bona fide component of the Gcn5-containing SAGA-type complex, increases the turnover of two shelterin proteins, TRF1 and POT1a (Atanassov, 2009). Our studies provided the first indication that Gcn5 and mammalian SAGA influence telomere maintenance and the first demonstration that SAGA affects protein stability. We are now defining the role of Gcn5 and SAGA in maintenance of pluripotency in ES cells and ES cell differentiation. Our data indicate that Gcn5 is an important cofactor for both Myc and E2F family transcription factors. We are also identifying additional USP22 protein substrates and are defining USP22 functions during mouse development.

We have also discovered unexpected functions for the Set1 lysine methyltransferase, which is highly homologous with the MLL protein, during mitosis. We found that Set1 methylates kinetochore proteins, such as Dam1, in yeast, and we have determined that H2Bub is required in trans for methylation of this protein. This work provides the first example of cross talk between a histone modification and a modification in a non-histone protein, defining a new mode of “chromatin signaling.”

Zhang K, Lin W, Latham JA, Riefler GM, Schumacher JM, Chan C, Tatchell K, Hawke DH, Kobayashi R, Dent SY. The Set1 methyltransferase opposes Ipl1 aurora kinase functions in chromosome segregation. Cell 122(5):723-34, 9/2005. PMCID: PMC1794220

Atanassov BS, Evrard YA, Multani AS, Zhang Z, Tora L, Devys D, Chang S, Dent SY. Gcn5 and SAGA Regulate Shelterin Protein Turnover and Telomere Maintenance. Mol Cell 35(3):352-364, 8/2009. PMCID: PMC2749492.

Atanassov BS, Dent SY. USP22 regulates cell proliferation by deubiquitinating the transcriptional regulator FBP1. EMBO Rep 12(9):924-30, 9/2011. e-Pub 9/2011. PMCID: PMC3166460.

Latham JA, Chosed RJ, Wang S, Dent SY. Chromatin signaling to kinetochores: Trans-regulation of Dam1 methylation by histone H2B ubiquitination. Cell 146(5):709-19, 9/2011. PMCID: PMC3168986.

Citation Format: Sharon Y.R. Dent, Boyko Atanassov, Calley Hirsch, Evangelia Koutelou, John Latham. New functions for histone modifying enzymes. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA07.
Abstract IA08: Targeting histone methylation in leukemia
Authors: Armstrong Scott A.
Published: August 2014

Leukemias harboring rearrangements of the MLL gene carry a poor prognosis. Over the past 6 years, it has become increasingly clear that fusions of MLL induce widespread epigenetic deregulation that may mediate much of their transforming activity. The histone methyltransferase DOT1L, which methylates histone 3 on lysine 79 (H3K79), has received particular attention. Genome-wide H3K79 methylation profiles in MLL-rearranged leukemias are abnormal, and can serve to distinguish MLL-rearranged from other types of leukemias. Loss of H3K79 methylation affects expression of MLL-target loci and is detrimental to the leukemogenic activity of MLL-rearranged cells, suggesting that a DOT1L dependent, aberrant epigenetic program drives transformation in these leukemias. Small molecule DOT1L inhibitors have been developed and selectively inhibit proliferation of MLL-rearranged leukemia cells. These molecules are now in clinical trials for patients with relapsed refractory leukemia. Also, the Polycomb Repressive Complex 2 (PRC2) has been implicated in self-renewal and cancer progression, and its components are overexpressed in many cancers, but its role in cancer development and progression remains unclear. We have used conditional alleles for the PRC2 components Enhancer of Zeste 2 (Ezh2) and Embryonic Ectoderm Development (Eed) to characterize the role of PRC2 function in leukemia development and progression. Compared to wildtype leukemia, Ezh2-null MLL-AF9-mediated acute myeloid leukemia (AML) show decreased organ infiltration and failed to accelerate upon secondary transplantation. However, Ezh2-null leukemias maintained self-renewal up to the third round of transplantation, indicating that Ezh2 is not strictly required for MLL-AF9 AML. In contrast, inactivation of the essential PRC2 gene, Eed led to complete ablation of PRC2-function, which is incompatible with leukemia growth. Gene expression array analyses indicated more profound gene expression changes in Eed-null compared to Ezh2-null leukemic cells, including downregulation of Myc target genes and upregulation of PRC2 targets. These data show that histone-modifying enzymes play critical roles in leukemia and may be relevant therapeutic targets in these diseases.

Citation Format: Scott A. Armstrong. Targeting histone methylation in leukemia. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA08.
Abstract IA15: Epigenetics of hormone dependence
Authors: Brown Myles
Published: August 2014

A detailed understanding of interplay between genetics and epigenetics in the progression and hormone dependence of distinct cancer subtypes is critical to the development of new approaches to prevention and therapy. Steroid hormones and their receptors play essential roles in the development and progression of both breast and prostate cancers. It remains unknown what role an epigenetically defined cell of origin plays in determining hormone responsiveness. In addition whether acquired resistance to endocrine therapy is the result of clonal evolution or the presence of hormone independent tumor-initiating cells, or both remains to be determined. Gain of function mutations and gene amplification of the androgen receptor (AR) in prostate cancer and the estrogen receptor (ER) in breast cancer in a subset of hormone refractory cases supports clonal evolution as the dominant mechanism of resistance in in these patients. We have investigated how the pathways downstream of AR and ER are activated in cases where they are not genetically altered and whether these depend on mutations in other epigenetic regulators including other transcription factors involved in steroid receptor networks and chromatin modifying enzymes involved in transcriptional regulation mediated by these receptors. Renewed success in targeting the steroid receptors themselves and promising advances in inhibiting the activity of the chromatin modifying enzymes provides new opportunities for the treatment of patients with breast and prostate cancer.

Citation Format: Myles Brown. Epigenetics of hormone dependence. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA15.
Abstract IA17: The stem cell factor LIN28 in pediatric cancer
Authors: Daley George Q.
Published: August 2014

LIN28, a developmental regulator originally identified as a heterochronic mutant in C. elegans, functions as an RNA binding protein that regulates protein expression through effects on mRNA translation and through control of processing of the let-7 family of tumor suppressor microRNAs. Lin28 has two paralogs, Lin28A and Lin28B, and studies have revealed a role for these factors in a wide array of biological phenotypes including organismal growth and metabolism, sexual maturation, and oncogenesis. LIN28/let-7 act antagonistically to regulate the balance of self-renewal and differentiation in embryonic stem cells, and LIN28 is one of the factors shown to reprogram somatic cells to pluripotency. In pediatric cancers, high level expression of LIN28 is a consistent marker of germ cell tumor, frequently observed in neuroblastoma and hepatoblastoma, and activated by chromosomal translocation in Wilms tumor. Thus, the stem cell factor LIN28 is commonly dysregulated in pediatric tumors and may represent a novel therapeutic target.

Citation Format: George Q. Daley. The stem cell factor LIN28 in pediatric cancer. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA17.
Abstract IA20: Epigenetic deregulation of gene expression
Authors: Melnick Ari
Published: August 2014

Epigenetic deregulation of gene expression through aberrant DNA methylation or histone modification plays an important role in the malignant transformation of hematopoietic cells. In particular, acute myeloid leukemias (AMLs) can be classified according to epigenetic signatures affecting DNA methylation or histone modifications affecting specific gene sets. Heterozygous somatic mutations in the loci encoding isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in ~20% of AMLs and are accompanied by global DNA hypermethylation and hypermethylation and silencing of a number of specific gene promoters. IDH1/2 mutations are almost completely mutually exclusive with somatic loss of function mutations in TET2, which hydroxylates methylcytosine (mCpG). DNA hydroxymethylation can function as an intermediate step in mCPG demethylation. TET2 mutant de novo AMLs also display global and promoter specific hypermethylation partially overlapping with IDH1/2 mutant cases. Mutations in the IDH1/2 loci result in a neomorphic enzyme that generates the aberrant oncometabolite 2-hydroxyglutarate (2HG) using alpha-ketoglutarate (aKG) as a substrate. 2HG can disrupt the activity of enzymes that use aKG as a cofactor, including TET2 and the Jumonji family of histone demethylases. Expression of mutant IDH isoforms inhibit TET2 hydroxymethylation and Jumonji histone demethylase functions. IDH and TET2 mutant AMLs accordingly exhibit reduced levels of hydroxymethylcytosine and a trend towards reduced histone methylation. Mutant IDH or TET2 loss of function causes differentiation blockade and expansion of hematopoietic stem cells and TET2 knockout results in a myeloproliferative phenotype in mice. Hydroxymethylcytosine is abundance in hematopoietic stem cells, and displays specific distribution patterns, and yet the function of this covalent modification is not fully understood. Recent data link TET2 with the function of cytosine deaminases as a pathway towards DNA demethylation, which has implications as well for B-cell lymphomas and CML lymphoid blast crisis, which are linked with the actions of activation induced cytosine deaminase. Altogether, the available data implicate mutations in IDH1/2 and TET2 in promoting malignant transformation in several tissues, by disrupting epigenomics programming and altering gene expression patterning.

Citation Format: Ari Melnick. Epigenetic deregulation of gene expression. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA20.
Abstract PR03: Multivalent histone engagement by the linked tandem Tudor and PHD domains of UHRF1 is required for DNA methylation maintenance
Authors: Rothbart Scott B. , Dickson Bradley M. , Ong Michelle S. , Krajewski Krzysztof , Houliston Scott , Kireev Dmitri B. , Arrowsmith Cheryl H. , Strahl Brian D.
Published: August 2014

DNA methylation and histone post-translational modifications (PTMs) represent two key epigenetic regulators of DNA information in chromatin. The faithful inheritance of DNA methylation is essential for normal mammalian development and long-term transcriptional silencing, and aberrant DNA methylation patterns are a hallmark of many cancers. Histone PTMs can alter the physical structure of chromatin and have been proposed to function in the context of a “histone code” to coordinate the recruitment of effector proteins that elicit selective effects on chromatin-templated cellular processes like gene transcription and DNA repair. Indeed, defects in histone recognition by effector proteins are also emerging as underlying causes of cancer initiation and progression. Effector proteins often contain multiple chromatin binding domains. While significant progress has been made characterizing individual effector domains, the role of paired domains and how they function in a combinatorial fashion within chromatin is poorly defined. The E3 ubiquitin ligase UHRF1 is a unique chromatin effector protein, in that it integrates the recognition of both histone PTMs and DNA methylation. We recently showed that H3K9 methyl binding by the tandem Tudor domain (TTD) of UHRF1 is required for DNA methylation maintenance in cancer cells — yet the function of its adjacent plant homeodomain (PHD) in this process was unknown. Here we show that the linked TTD and PHD of UHRF1 operate as a single functional unit in cells, providing a defined combinatorial readout of the modification state of the histone H3 tail that is essential for UHRF1-directed DNA methylation maintenance by DNMT1. These findings provide critical support for the ‘histone code' hypothesis, demonstrating that multivalent histone engagement plays a fundamental role in driving a downstream biological event in chromatin and suggest UHRF1 may be a favorable therapeutic target for regulating aberrant DNA methylation in cancers.

This abstract is also presented as Poster B26.

Citation Format: Scott B. Rothbart, Bradley M. Dickson, Michelle S. Ong, Krzysztof Krajewski, Scott Houliston, Dmitri B. Kireev, Cheryl H. Arrowsmith, Brian D. Strahl. Multivalent histone engagement by the linked tandem Tudor and PHD domains of UHRF1 is required for DNA methylation maintenance. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr PR03.
Abstract PR12: CHD5 is required for neurogenesis and has a dual role in facilitating gene expression and Polycomb gene repression
Authors: Egan Christopher M. , Nyman Ulrika , Skotte Julie , Streubel Gundula , Turner Siobhán , O'Connell David J. , Rraklli Vilma , Dolan Michael J. , Chadderton Naomi , Klaus Hansen , Jane Farrar Gwyneth , Helin Kristian , Holmberg Johan , Bracken Adrian P.
Published: August 2014

The chromatin remodeler CHD5 is expressed in neural tissues and is frequently deleted in aggressive neuroblastoma. Very little is known about the normal function of CHD5, its mechanism of action or how its deletion may contribute to neuroblastoma. Here we report that depletion of CHD5 in the developing murine neocortex blocks neuronal differentiation leading to an accumulation of undifferentiated progenitors. Consistent with this, ChIP-SEQ analysis of CHD5 shows it binds a large cohort of neuronal genes and is required for facilitating their activation during differentiation. However, CHD5 also binds a cohort of Polycomb target genes and is required for the maintenance of H3K27me3 on their promoters and their sustained repression during differentiation. Finally, we show that the chromodomains of CHD5 directly bind H3K27me3 and are essential for the function of CHD5 during neuronal differentiation. These findings provide new insights into the essential role of CHD5 during neurogenesis and show how the inactivation of this candidate tumor suppressor might contribute to the progression of neuroblastoma via a defect in differentiation.

This abstract is also presented as Poster A05.

Citation Format: Christopher M. Egan, Ulrika Nyman, Julie Skotte, Gundula Streubel, Siobhán Turner, David J. O'Connell, Vilma Rraklli, Michael J. Dolan, Naomi Chadderton, Hansen Klaus, Farrar Gwyneth Jane, Kristian Helin, Johan Holmberg, Adrian P. Bracken. CHD5 is required for neurogenesis and has a dual role in facilitating gene expression and Polycomb gene repression. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr PR12.
Abstract PR16: A new class of small molecule acetyltransferase inhibitors discovered through high-throughput screening are potent anticancer agents with cancer-type specific activity
Authors: Yang Heng , Pinello Christie E. , Luo Jian , Li Dawei , Wang Yunfei , Lisa Zhao Y. , Jahn Stephan C. , Saldanha S. Adrian , Planck Jamie , Geary Kyla R. , Ma Haiching , Law Brian K. , Roush William R. , Hodder Peter , Liao Daiqing
Published: August 2014

Acetyltransferase p300 (KAT3B) plays key roles in signaling cascades that support cancer cell survival and sustained proliferation. p300 is overexpressed in several types of cancer, which correlates with tumor progression. Thus, p300 represents a rational anticancer therapeutic target. To discover novel anticancer agents that target p300, we conducted a high-throughput screening campaign. A library of 622,079 compounds was assayed for cytotoxicity to the triple-negative breast cancer (TNBC) cell line MDA-MB-231 but not to the human mammary epithelial cells. The resulting compounds were tested in a biochemical assay for inhibiting the enzymatic activity of p300. One compound (L002, NSC764414) displayed an IC50 of 1.98 μM against p300 in vitro, inhibited acetylation of histones and p53, and suppressed STAT3 activation in cell-based assays. L002 could be docked to the active site of the p300 catalytic domain. Biochemical tests of a series of related compounds revealed functional groups that may impact inhibitory potency of L002 against p300. Interestingly, these analogs showed inhibitory activities against CBP (the cellular paralog of p300), PCAF and GCN5, but not to other acetyltransferases (KAT5, KAT6B and KAT7), histone deacetylases (HDACs) and histone methyltransferases. Among the NCI-60 panel of cancer cell lines, leukemia and lymphoma cell lines were extremely sensitive to L002, whereas it is toxic to only a limited number of cell lines derived from solid tumors. Notably, breast cancer cell lines, especially those derived from TNBC, were highly susceptible to L002. In vivo, it potently suppressed tumor growth and histone acetylation of MDA-MB-468 xenografts. Thus, these new acetyltransferase inhibitors are potential anticancer therapeutics with specific activities against breast cancer and hematological malignancies.

This abstract is also presented as Poster B43.

Citation Format: Heng Yang, Christie E. Pinello, Jian Luo, Dawei Li, Yunfei Wang, Zhao Y. Lisa, Stephan C. Jahn, S. Adrian Saldanha, Jamie Planck, Kyla R. Geary, Haiching Ma, Brian K. Law, William R. Roush, Peter Hodder, Daiqing Liao. A new class of small molecule acetyltransferase inhibitors discovered through high-throughput screening are potent anticancer agents with cancer-type specific activity. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr PR16.
Abstract A23: Novel ursolic acid derivatives with potent antitumor activity
Authors: Leal Ana S. , Salvador Jorge A.R. , Yongkui Jing
Published: August 2014

Cancer is the fourth leading cause of death worldwide, within pancreatic cancer is the fourth leading cause of cancer death and without effective medical therapy.1 Natural products provide a promising source for developing effective anticancer agents.2 Ursolic acid is a pentacyclic triterpenoid present in plants, vegetables, and fruits that has been found to have several biological activities, including antitumor activity.3 In order to improve the preexisting antitumor activity of ursolic acid, a panel of semisynthetic derivatives were prepared, characterized, and evaluated for the antiproliferative activities in several cancer cell lines.

Ursolic acid was chemically modified by the introduction of heterocyclic rings in several points of the backbone structure and was found that modifications at C2 had improved activities against proliferation of pancreatic cancer cell lines, inducing apoptosis with upregulation of p53 and NOXA levels, and downregulation of XIAP levels.

Fluor is a highly desirable atom, in key positions of active molecules can improve metabolic and chemical stability, membrane permeability and binding affinity.4 We have introduced fluor into the ring C of ursolic acid and a series of ursane-type fluor-derivatives were synthesized. The most potent derivative was 19-fold more active than ursolic acid to inhibit Aspc-1 cell growth and to arrest cell cycle at G1 phase with upregulation of p21waf1, and apoptosis at higher concentrations with upregulation of NOXA and downregulation of c-FLIP.

These new semisynthetic derivatives of ursolic acid may have potential for drug discovery to treat cancer.

Citation Format: Ana S. Leal, Jorge A.R. Salvador, Jing Yongkui. Novel ursolic acid derivatives with potent antitumor activity [abstract]. In: Proceedings of the AACR Special Conference on Chemical Systems Biology: Assembling and Interrogating Computational Models of the Cancer Cell by Chemical Perturbations; 2012 Jun 27-30; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2012;72(13 Suppl):Abstract nr A23.
Abstract A10: In silico design of biomarker-optimized drug combinations in ERBB2+ cancers
Authors: Du Jinyan , Kirouac Daniel , Lahdenranta Johanna , Overland Ryan , Onsum Matthew , McDonagh Charlotte
Published: August 2014

Crosstalk and compensatory circuits within cancer signaling networks fundamentally limit the activity of targeted therapies. Combination drug regimens are thus required to fully inhibit an oncogenic network. However, rationally designing optimal combinations, given the large number of targeted agents available remains a challenge. Previously, we developed a multi-scale systems model of ERBB2-high breast cancer to quantitatively interrogate relationships between biomarkers (proxies for the mode of network activation) and combination drug efficacy. We found that targeting the ERBB2-ERBB3 heterodimer with a combination of ERBB2 inhibitors (trastuzumab and lapatinib) and an ERBB3 inhibitor (MM-111) was more effective at inducing tumor regression than the combination of an AKT and MEK inhibitor (MK2206 and GSK1120212), and was significantly better tolerated. However, the model was parameterized based on data collected from one cell line. Here we have performed profiling of an 18-cell line panel to assess whether our model and results are extendable to other ERBB2-high cancers. We monitored cell viability and signaling events upon AKT and MEK inhibitor treatments in the presence or absence of the ERBB3 ligand heregulin in these cell lines. While all cells were ERBB2+, we observed widely variable phenotypic and signaling dependencies on the PI3K/AKT and MAPK/ERK pathway activities across the panel. At the phenotypic level, cells primarily depend on either AKT or ERK signaling in basal conditions. Interestingly, upon heregulin stimulation some cells lines switch pathway dependency from AKT to ERK. Adaptive feedback circuits downstream of ERK and AKT were identified in all cell lines, though the identity and strength vary extensively. ERBB3 signaling and total AKT were consistently up-regulated to various degrees upon AKT inhibitor treatment. In contrast, multiple ERBB receptors as well as other RTKS, AKT signaling, and total ERK were upregulated in the response to MEK inhibition. While it is plausible that signaling pathways beyond AKT and ERK are modulating cell viability, we are able to quantitatively describe cell growth regulation based on AKT and ERK pathway activity using quantitative logic-based modeling framework. Our results will help us better understand how signaling events are decoded by cancer cells into phenotypic responses, and enable in silico drug combination screening across molecularly and functionally heterogeneous cancers.

Citation Format: Jinyan Du, Daniel Kirouac, Johanna Lahdenranta, Ryan Overland, Matthew Onsum, Charlotte McDonagh. In silico design of biomarker-optimized drug combinations in ERBB2+ cancers. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A10.
Abstract A12: Combined EZH2 and PI3K inhibition as a novel therapy for treatment of castration resistant prostate cancer
Authors: Enos Miriam
Published: August 2014

Prostate cancer is the most frequently diagnosed non-dermatological malignancy in North American men. Presently, preventative testing and castration therapies are the only measures that effectively decrease mortality. However, these therapies are not effective in the case of the advanced disease, for which there is still no cure. As such, there is an unmet medical need to develop more effective therapies for castration-resistant, metastatic tumors.

Surprisingly, there are but a few things known about the signals that drive and maintain advanced disease. The PI3K/AKT signaling pathway is frequently deregulated in human malignancies and is activated in a majority of advanced stage carcinomas of the prostate. More recently, a member of PRC2 complex, EZH2, has been reported to be a marker and driver of late stage disease. Based on this knowledge about the key disregulated signaling pathways in prostate cancer, we decided to explore them as potential therapeutic avenues.

We found that both genetic and biochemical inhibition of EZH2 results in reduced proliferation and transformation in soft agar. Similarly, treating cells with a PI3K specific inhibitor, GDC- 0941 produces only a cytostatic effect, an observation that is supported by a body of literature suggesting that GDC-0941 is ineffective as a mono-therapy drug.

To this end we decided to explore the combination of both EZH2 and PI3K inhibitors and have found that simultaneous suppression of these two important oncogenic signals, kills androgen-dependent (AD) and independent (AI) prostate cancer cells in vitro and in vivo. Knock-down of another member of PRC2 complex, SUZ12, phenocopies this cytotoxicity when combined with PI3K inhibitor, suggesting that EZH2 role is PRC2-dependent.

These preliminary data suggest specific cooperation of EZH2 and PI3K/AKT signaling pathways in assuring cell survival and maintenance of tumorigenicity in prostate cancer cells. Currently we are exploring the potential downstream effectors that could be responsible for the synergy that we observe upon combination treatment.

Citation Format: Miriam Enos. Combined EZH2 and PI3K inhibition as a novel therapy for treatment of castration resistant prostate cancer. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A12.
Abstract A18: CDK6 binds and promotes the degradation of the EYA2 protein
Authors: Kohrt Dawn , Crary Jennifer , Zimmer Marc , Ford Heide , Hinds Phi , Grossel Martha
Published: August 2014

Cyclin dependent kinase 6 (Cdk6) is a D-cyclin activated kinase that is directly involved in driving the cell cycle through inactivation of pRB in G1 phase. Increasingly, evidence suggests that CDK6, while directly driving the cell cycle, may only be essential for proliferation of specialized cell types, agreeing with the notion that CDK6 also plays an important role in differentiation. Here, evidence is presented that CDK6 binds to and promotes degradation of the EYA2 protein. The EYA proteins are a family of proteins that activate genes essential for the development of multiple organs, regulate cell proliferation, and are misregulated in several types of cancer. This interaction suggests that CDK6 regulates EYA2 activity, a mechanism that could be important in development and in cancer.

Citation Format: Dawn Kohrt, Jennifer Crary, Marc Zimmer, Heide Ford, Phi Hinds, Martha Grossel. CDK6 binds and promotes the degradation of the EYA2 protein. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A18.
Abstract A21: Direct transcriptional functions of the APC tumor suppressor in regulating gene expression
Authors: Hankey William , McIlhatton Michael A. , Lan Xun , Jegga Anil G. , Jin Victor X. , Aronow Bruce J. , Groden Joanna
Published: August 2014

Mutation of the APC tumor suppressor gene occurs in approximately 75% of colorectal cancers (CRC) and is considered the central event driving tumor initiation in the large intestine. The APC protein controls gene expression indirectly through the canonical WNT signaling pathway as a component of a cytoplasmic complex that promotes the proteolysis of the transcriptional co-regulator β-catenin. Recent studies have shown that APC is also a chromatin-associated protein that interacts with the promoter of the MYC proto-oncogene to downregulate its transcription. We have tested the hypothesis that APC similarly regulates other genes relevant to tumorigenesis, and that the 75% of CRCs that are deficient in APC may show abnormal activation or suppression of these genes, in comparison to the other 25% of CRCs. These differences could potentially underlie differences in tumor progression and/or responses to chemotherapy, and could enable the identification of new therapeutic targets.

The APC genotype of two CRC cell lines was manipulated using siRNA to reduce APC expression in one cell line wild-type for APC and using an APC transgene to induce exogenous APC expression in a second cell line mutated for APC. NanoString and RNA-seq technologies have generated gene expression datasets from these cell lines and identify more than 700 genes that could be directly regulated at the transcriptional level by APC. RT-PCR and in silico analyses have validated a smaller subset of these genes. Functional pathway analysis has led to the hypothesis that APC loss facilitates evasion of apoptosis, with a subset of 17 pro- and anti-apoptotic candidate genes implicated in this effect. Critical experiments in progress will confirm the extent to which the aberrant expression of these genes in vivo is specific to colon tumors lacking APC, using Apc-deficient and Apc-wild-type mouse models of intestinal cancer, and will identify direct chromatin binding sites of APC by ChIP-seq. These approaches will validate our gene expression analyses and identify a set of direct gene targets that APC regulates.

Citation Format: William Hankey, Michael A. McIlhatton, Xun Lan, Anil G. Jegga, Victor X. Jin, Bruce J. Aronow, Joanna Groden. Direct transcriptional functions of the APC tumor suppressor in regulating gene expression. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A21.
Abstract A22: Mechanisms of oncogenic cooperation between EGFR/Ras and Hippo pathways in Drosophila and human cellular transformation models
Authors: Hong Xin , Cohen Stephen Michael
Published: August 2014

The hippo/Salvador kinase cascade is a highly conserved signaling pathway regulating growth and organ size from Drosophila to mammals. The invertebrate-specific microRNA, bantam, is a key target and effector downstream of the Drosophila hippo pathway. Bantam inhibits the pro-apoptotic factor hid and promotes growth through an as yet unidentified mechanism. The EGFR/Ras pathway coordinates growth and patterning in Drosophila. We characterize a molecular circuit involving bantam, socs36E and EGFR that regulates tissue growth. Socs36E is a direct bantam target that negatively regulates wing size. Socs36E buffers EGFR activity by acting in a negative feedback loop with EGFR. Interestingly, bantam and socs36E repress each other forming a potential bistable switch. This switch might allow effective repression of socs36E during the rapid growth phase of wing disc development. Therefore, bantam represents a unique molecular linkage connecting hippo and EGFR signaling in promoting growth. Furthermore, we found that the mammalian socs family members SOCS5 and SOCS6 are targeted by an oncogenic miRNA, which plays a role in RasV12 mediated tumoriogenesis in soft agar assays. Therefore, the SOCS family proteins are evolutionarily conserved regulators of growth and cancer transformation.

Citation Format: Xin Hong, Stephen Michael Cohen. Mechanisms of oncogenic cooperation between EGFR/Ras and Hippo pathways in Drosophila and human cellular transformation models. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A22.
Abstract A30: Dysregulation of BMI1 and microRNA-16 collaborate to enhance an anti-apoptotic potential in the side population of refractory mantle cell lymphoma
Authors: Kitadate Akihiro , Teshima Kazuaki , Ohshima Kouichi , Seto Masao , Sawada Kenichi , Tagawa Hiroyuki
Published: August 2014

The proto-oncogene BMI1 and its product, Bmi1, is overexpressed in various types of tumors, particularly in aggressive tumors and tumors resistant to conventional chemotherapy. BMI1/Bmi1 is also crucially involved in cancer-initiating cell maintenance, and is recurrently upregulated in mantle cell lymphoma (MCL), especially aggressive variants. Recently, side population (SP) cells were shown to exhibit tumor-initiating characteristics in various types of tumors. In this study, we show that recurrent MCL cases significantly exhibit upregulation of BMI1/Bmi1. We further demonstrate that clonogenic MCL SP shows such tumor-initiating characteristics as high tumorigenicity and self-renewal capability, and that BMI1 was upregulated in the SP from recurrent MCL cases and MCL cell lines. On screening for upstream regulators of BMI1, we found that expression of microRNA-16 (miR-16) was downregulated in MCL SP cells by regulating Bmi1 in the SPs, leading to reductions in tumor size following lymphoma xenografts. Moreover, to investigate downstream targets of BMI1 in MCL, we performed cross-linking/chromatin immunoprecipitation assay against MCL cell lines and demonstrated that Bmi1 directly regulated pro-apoptotic genes such as BCL2L11/Bim and PMAIP1/Noxa, leading to enhance anti-apoptotic potential of MCL. Finally, we found that a proteasome inhibitor bortezomib, which has been recently used for relapsed MCL, effectively induced apoptosis among MCL cells while reducing expression of Bmi1 and increasing miR-16 in MCL SP. These results suggest that upregulation of BMI1 and downregulation of miR-16 in MCL SP has a key role in the disease's progression by reducing MCL cell apoptosis. Our results provide important new insight into the pathogenesis of MCL and strongly suggest that targeting BMI1/Bmi1 might be an effective approach to treating MCL, particularly refractory and recurrent cases.

Citation Format: Akihiro Kitadate, Kazuaki Teshima, Kouichi Ohshima, Masao Seto, Kenichi Sawada, Hiroyuki Tagawa. Dysregulation of BMI1 and microRNA-16 collaborate to enhance an anti-apoptotic potential in the side population of refractory mantle cell lymphoma. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A30.
Abstract A37: MicroRNA-26b potentiates cytotoxicity of malignant glioma cells by radiation
Authors: Lee David J. , Hwang Yeo-Hyun , Kim In Ah
Published: August 2014

MicroRNA-26b is known to be upregulated in high grade glioma and to play a crucial role in epithelial to mesenchymal transition (EMT). In this study, we evaluated the potential of targeting miRNA-26b to overcome the radioresistance of human malignant glioma cells. We also aimed to elucidate the mechanisms of radiosensitization by miR-26b.

U251 human glioma cells having activation of PI3K signaling due to deletion of PTEN were transfected with pre-miR-26b or control pre-miRNA. Ectopic overexpression of miR-26b attenuated the expression of p-AKT and enhanced the cytotoxicity of U251 cells by radiation. Overexpression of miR-26b prolonged radiation-induced gamma H2AX foci formation and attenuated the expression of p-DNA-PKcs. Increased expression of mature miR-26b also suppressed invasion, migration, vascular tube formation, and led to down-regulation of HIF-1alpha, VEGF and MMP-2 expression. In contrast, overexpression of miR-26b did not increased radiaton toxicity of normal human astrocytes.

Taken together, these findings suggest that miR-26b could be an attractive target for overcoming radioresistnace of human malignant glioma cells via unique modulation of the cancer pro-survival signaling and EMT.

Citation Format: David J. Lee, Yeo-Hyun Hwang, In Ah Kim. MicroRNA-26b potentiates cytotoxicity of malignant glioma cells by radiation. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr A37.
Abstract B22: Modulation of insulin-like growth factor binding proteins expression in human prostate cancer cells in vitro by american cranberry (Vaccinium macrocarpon) extract
Authors: Hurta Robert , McKeown Brendan , Boddeti Ravi , Neto Catherine
Published: August 2014

American cranberry (Vaccinium macrocarpon) extract has been previously shown to affect cell cycle, apoptosis, and proliferative ability in prostate cancer cells in vitro. The insulin-like growth factors (IGFs) and their modulators the insulin-like growth factor binding proteins (IGFBPs) may play roles in these events. IGFBPs act by regulating IGF action but also exhibit many IGF-independent activities. IGFBP expression in response to cranberry extract [50 & 100 ug/mL for 3 and 24 hours] was determined via Western blot analysis in DU145 and in PC3 androgen-refractory human prostate cancer cells. Cranberry extract treatment decreased the expression levels of both IGFBP-2 and IGFBP-5 and increased the expression levels of IGFBP-4. IGFBP-3 protein expression levels were apparently unaffected by cranberry extract suggesting that IGFBP-3 is likely not involved in this cellular response to cranberry extract. A reduction in IGFBP-2/-5 expression suggests a reduction in IGF through loss of IGF-stabilization by the IGFBPs suggesting an inhibition of the cell cycle due to this restriction in IGF. Increases in IGFBP-4 levels in response to cranberry would also inhibit IGFs and thereby inhibit cellular growth and proliferation. Increased IGFBP-2/-5 expression has been associated with progression from androgen-sensitivity to androgen-insensitivity in prostate cancer cells. Decreased IGFBP-2/-5 expression levels in response to cranberry extract could counteract this progression in vitro. [funded by The Cranberry Institute (Wisconsin Board) and Jeanne & J.-Louis Levesque Foundation]

Citation Format: Robert Hurta, Brendan McKeown, Ravi Boddeti, Catherine Neto. Modulation of insulin-like growth factor binding proteins expression in human prostate cancer cells in vitro by american cranberry (Vaccinium macrocarpon) extract. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr B22.