bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1046: The role of NF-kB activation in the immortalization of nasopharyngeal epithelial cells
Authors: Tsao George Sai-Wah , Zhu Dan Dan , Zhang Jun , Deng Wen
Published: August 2015

NF-κB is a key regulator of inflammatory response and is frequently activated in human cancers including nasopharyngeal carcinoma (NPC), which is closely associated with Epstein-Barr virus (EBV) infection and a common cancer in Southern China including Hong Kong. However, it is well-established that EBV infection alone is not sufficient for NPC development. Other genetic alterations in host cells and the interaction between EBV infection and host cell microenvironment are required for NPC development. Our group has been involved in illustrating molecular events underlying the immortalization of nasopharyngeal epithelial (NPE) cells. Molecular changes which facilitate immortalization may represent early genetic events in human carcinogenesis. We found that the activation of NF-κB was a common feature in telomerase-immortalized NPE cell lines. We further showed that canonical NF-κB activation pathway was involved in the immortalization of NPE cells. The activation of NF-κB was essential for the proliferation of the immortalized NPE cells because suppression of NF-κB activity could selectively inhibit the proliferation of immortalized but not primary NPE cells. The upregulation of c-Myc and Bmi-1, known to promote cell proliferation, were associated with NF-κB activation in the immortalized NPE cell lines. Interestingly, mTOR was found to be the upstream activator of NF-κB. We further showed that EGFR/Erk(1/2) pathway was the upstream regulator of mTOR/NF-κB pathway, revealing the pathway of EGFR/Erk(1/2)/mTOR/NF-κB for NF-κB activation in telomerase-immortalized NPE cells. Elucidation of specific pathway leading to NF-κB activation in immortalized NPE cells may open up opportunity for early intervention to suppress or prevent NPC development.

Citation Format: George Sai-Wah Tsao, Dan Dan Zhu, Jun Zhang, Wen Deng. The role of NF-kB activation in the immortalization of nasopharyngeal epithelial cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1046. doi:10.1158/1538-7445.AM2015-1046
Abstract 105: A rational approach for the discovery of inhibitors of NSD2 for the treatment of cancer
Authors: Fromond Claudia , Espanel Xavier , Soudé Anne , Chene Laurent , Masson Philippe , Boubia Benaïssa , Montalbetti Christian , Broqua Pierre
Published: August 2015

Multiple myeloma (MM) is a plasma cell malignancy which accounts for approximately 10% of hematologic malignancies. Despite the introduction of new therapeutic agents, MM remains incurable and nearly all patients ultimately relapse. About 20% of MM are due to a chromosomal translocation t(4;14) leading to overexpression of the NSD2 histone methyltransferase. NSD2 catalyzes dimethylation of lysine 36 on histone H3 (H3K36me2) and is associated with transcriptionally active regions. Several studies have shown that in MM harboring the translocation t(4;14), oncogenic programming is dependent on the methyltransferase activity of NSD2. In addition, the NSD2 overactivity is also observed in prostate and lung cancers. Thus, NSD2 is a potential target for cancers, for which no selective drug is available to date.

Using the AlphaLisa™ technology, we screened 240,000 compounds coming from our proprietary library. The assay is based on the detection of H3K36me2 on nucleosome by a specific antibody. False positives hits were removed by a technological counterscreen, and compounds with redox activity were excluded. This strategy allowed us to identify about 200 compounds. To focus on the most interesting ones, an orthogonal counterscreen based on 3H SAM incorporation is being completed. In parallel to the biochemical screening, we are developing secondary cellular assays based on the H3K36me2 methylation and proliferation to further confirm hit activity. To our knowledge, no NSD2 inhibitor have been identified to date despite several screening effort performed by other groups. Our library has already produced new chemical starting points for other KMTs, and we believe that our hits could be promising starting points to generate potent and selective NSD2 inhibitors.

Citation Format: Claudia Fromond, Xavier Espanel, Anne Soudé, Laurent Chene, Philippe Masson, Benaïssa Boubia, Christian Montalbetti, Pierre Broqua. A rational approach for the discovery of inhibitors of NSD2 for the treatment of cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 105. doi:10.1158/1538-7445.AM2015-105
Abstract 1050: Effects of 5-hydroxymethylcytosine epigenetic modification on G-quadruplex and i-motif structure and stability within the VEGF promoter
Authors: Morgan Rhianna , Wadkins Randy , Brooks Tracy
Published: August 2015

CpG islands are prone to epigenetic cytosine modification via methylation (5mC) or 5-hydroxymethylation (5hmC). Although their potential sequences and regions for occurrence in G/C-rich DNA overlap, throughout the genome 5mC and 5hmC, and 5mC and G-quadruplex (G4) and i-motif (iM) structures, are found to not be co-existent. Moreover, in some regions of DNA, 5hmCs and G4/iM forming regions have high potential to form in concurrence. However, minimal research has been done examining the effect of 5hmC incorporation into G4 and iMs in order to determine the effect of cytosine modification on non-B-DNA structures. These higher order secondary DNA formations have been described within the VEGF promoter and have great potential to silence its transcription and prevent subsequent angiogenesis. In addition, there are three cytosines in the promoter on both the guanine- and the cytosine-rich strands found in CpG sequences that have the potential for 5mhC modification. Therefore, the current study was undertaken to examine the whether there is an effect of 5hmC incorporation into G4 loops and iM stems on structure formation and stabilization. Significant changes in structure or stability in buffers with varying pH’s, cationic strength, and molecular crowding conditions were examined by circular dichroism and confirmed by UV-monitored melting. Electromobility shift assays differentiated inter- and intra-molecular structures, further identifying the number of isoforms. Notable changes in G4 dynamics were seen with two of the three loop modifications, which decreased overall stability and increased intermolecular structures. Much less marked effects were discovered for the iM structures. Ongoing studies examining the role of G4 and iM 5hmC incorporation in regulation by endogenous proteins are part of a global effort to best understand the physiological conditions regulating G4 and iM stability and function. These VEGF promoter higher order structures are promising therapeutic targets for anti-angiogenic therapy, and a comprehensive understanding of the physiological principles governing their formation will best inform future drug discovery efforts.

Brooks Lab startup funds, R15, University of Mississippi School of Pharmacy

Citation Format: Rhianna Morgan, Randy Wadkins, Tracy Brooks. Effects of 5-hydroxymethylcytosine epigenetic modification on G-quadruplex and i-motif structure and stability within the VEGF promoter. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1050. doi:10.1158/1538-7445.AM2015-1050
Abstract 1055: Genome-wide DNA methylation analysis in precancerous conditions associated with hepatitis B virus and hepatitis C virus infection
Authors: Arai Eri , Tian Ying , Gotoh Masahiro , Takahashi Yoriko , Ojima Hidenori , Kosuge Tomoo , Kanai Yae
Published: August 2015

The aim of this study is to reveal the significance of DNA methylation alterations in precancerous conditions associated with hepatitis virus infection. Genome-wide DNA methylation analysis using Infinium HumanMethylation450 BeadChip (Illumina) has been performed in 36 specimens of normal liver tissue obtained from patients without hepatocellular carcinomas (HCCs), 12 and 24 specimens of noncancerous liver tissues obtained from hepatitis B virus (HBV) infection-positive and hepatitis C virus (HCV) infection-positive patients with HCCs, respectively, and 24 samples of HCC themselves. On 21,106 CpG sites, alterations of DNA methylation levels were already evident in tissue specimens showing chronic hepatitis and liver cirrhosis, which are considered to be precancerous conditions, compared to normal liver tissues, and were inherited by HCC themselves. Unsupervised hierarchical clustering analysis based on DNA methylation levels at the 21,106CpG sites appropriately classified tissue specimens into normal, precancerous and cancerous clusters, indicating that DNA methylation alterations may participate continuously in hepatocarcinogenesis from the precancerous stage until cancers have become established. The incidence of DNA methylation alterations was higher at the precancerous stage of HCV-positive patients than that of HBV-positive patients. The incidence of DNA methylation alterations that actually occurred at the chronic hepatitis and liver cirrhosis stages but were restored in HCC themselves, which may be associated with inflammation or fibrosis, but may not participate in carcinogenesis itself, was at the almost equal levels regardless of probe annotations or hepatitis viruses. Many Infinium probes showing DNA methylation alterations participating continuously in multistage hepatocarcinogenesis were shared between HBV-positive and HCV-positive patients: only a small number of probes showed HBV-positive patient-specific DNA methylation alterations. On the other hand, DNA hypermethylation on regions around the transcription start sites was especially frequent during carcinogenesis in HCV-positive patients. We now intend to make the carcinogenetic risk estimation based on DNA methylation profiles at the precancerous stage clinically practical using liver biopsy specimens obtained prior to interferon therapy from patients who are followed up because of chronic liver diseases.

Citation Format: Eri Arai, Ying Tian, Masahiro Gotoh, Yoriko Takahashi, Hidenori Ojima, Tomoo Kosuge, Yae Kanai. Genome-wide DNA methylation analysis in precancerous conditions associated with hepatitis B virus and hepatitis C virus infection. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1055. doi:10.1158/1538-7445.AM2015-1055
Abstract 1062: Hypermethylation of BARHL2 gene is useful as a molecular marker in early stage of gastric cancer
Authors: Watanabe Yoshiyuki , Oikawa Ritsuko , Yamamoto Hiroyuki , Itoh Fumio
Published: August 2015

Aim: Although minimal invasive treatment is accepted for gastric cancer, we still do not have any appropriate risk markers to detect residual neoplasia and recurrence. We previously reported that aberrant DNA methylation is an early and frequent process in gastric carcinogenesis and could be useful for detection of neoplasia. Our goal is to find some candidate genes as a treatment marker using genome wide DNA methylation analysis for early gastric cancer (EGC). Methods and Results: We studied Methylated CpG Island Amplification Microarray (MCAM) analysis using 12 gastric washes (each 6 before (Pre) and after (Post) endoscopic treatment in same patients). One of them is BARHL2. We examined the DNA methylation status of BARHL2 in a validation set consisting of 128 washes samples (Pre, 64: Post, 64) at EGC. We next identified functional studies about BARHL2 as a putative tumor suppressor gene. Treating GCa cells that lacked BARHL2 expression with a methyltransferase inhibitor, 5-aza-2′-deoxycytidine, restored the gene's expression. Introduction of exogenous BARHL2 into silenced cells suppressed colony formation. Moreover, BARHL2 showed significantly differential methylation between before and after treatment in EGC patient (BARHL2, p<0.0001). Discussion: Our data suggest that silencing of BARHL2 occurs frequently in EGC and may play a key role in development and progression of the disease.

Citation Format: Yoshiyuki Watanabe, Ritsuko Oikawa, Hiroyuki Yamamoto, Fumio Itoh. Hypermethylation of BARHL2 gene is useful as a molecular marker in early stage of gastric cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1062. doi:10.1158/1538-7445.AM2015-1062
Abstract 1065: RASSF- 1A methylation in squamous cell carcinoma of the lower lip using pyrosequencing technique
Authors: Kaminagakura Estela , Coutinho-Camillo Cláudia , Strieder Luciana , Perez Danyel , Soares Fernando , Oliveira Kátia , Kowalski Luiz Paulo
Published: August 2015

Background: Chronic exposure to UVB radiation can cause hypermethylation and silencing of the tumor suppressor gene RASSF-1A. The protein encoded by this gene has multiple important functions in cell cycle control. Objective: to investigate the clinical significance of the methylation profile of RASSF-1A and of its protein expression in lower lip squamous cell carcinoma (SCC). Methods: The demographic and clinical-pathological data of 87 patients with a confirmed diagnosis were collected from the medical records. DNA was extracted from paraffin-embedded tissue samples and treated with sodium bisulfite. 2ug of treated and intact DNA was submitted to PCR amplification using a specific primer targeting the RASSF-1A promoter segments 1 and 2. RASSF1A methylation was investigated by pyrosequencing; the complementary strand was synthesized automatically and the signal emitted during the reaction was detected with a camera and transformed into peaks on a pyrogram. Results: Lower lip SCC was more frequent in men than women, with a proportion of 2.4:1. The mean patient age was 63.3 years. Lymph node involvement was absent (N0) in 73 (83.1%) of cases and present (N+) in 14 (16.1%). In group N0, most patients (n = 49, 67.1%) were in an early clinical stage (CS I and II). Analysis of the methylation of RASSF1A_01 was possible in 29 of the SCC cases. Hypermethylation was observed in 15 (51.7%) cases, one (3.4%) of them being associated with metastasis to regional lymph nodes. Analysis of RASSF1A_02 was possible in 20 cases and hypermethylation was observed in only 9 (45.0%). No significant correlation was found between the clinical-pathological parameters (age, gender, clinical stage) and aberrant methylation of the promoter region studied (p>0.05). Conclusions: Hypermethylation of the two RASSF-1A segments studied was observed in approximately 50% of SCC cases, but was not correlated to clinicopathologic parameters.

Citation Format: Estela Kaminagakura, Cláudia Coutinho-Camillo, Luciana Strieder, Danyel Perez, Fernando Soares, Kátia Oliveira, Luiz Paulo Kowalski. RASSF- 1A methylation in squamous cell carcinoma of the lower lip using pyrosequencing technique. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1065. doi:10.1158/1538-7445.AM2015-1065
Abstract 1067: Characterization of DNA hypermethylation in two cases of peritoneal mesothelioma
Authors: Oikawa Ritsuko , Watanabe Yoshiyuki , Hama Ryota , Yamamoto Hiroyuki , Itoh Fumio
Published: August 2015

Background: Malignant mesothelioma (MM) is a rare disease with a poor prognosis. Pleural mesothelioma, which is the most common type of MM, is considered to be caused by asbestos exposure and is increasing in incidence, with about 15,000 new cases diagnosed worldwide annually. On the other hand, peritoneal mesothelioma is a very rare type of MM; thus its pathogenesis is even less understood than pleural mesothelioma. Recent research on the pathogenesis of malignant pleural mesothelioma has indicated that both epigenetic and genetic alterations contribute to tumorigenesis. Here, we hypothesize that peritoneal mesothelioma also has an epigenetic alteration in the same genes (KAZALD1, TMEM30B and MAPK13). Our goal is to identify DNA methylation of these three candidate genes in two peritoneal mesothelioma cases. Methods: Laser capture microdissection (LCM) was used to separate diseased sections of formalin-fixed paraffin-embedded (FFPE) samples from one surgically resected tissue (epithelial type) and one autopsy tissue (sarcomatous type). Genomic DNA was subsequently extracted by the standard phenol chloroform method. The DNA was then treated with sodium bisulfite, and pyrosequencing analysis was used to quantitatively analyze the methylation of candidate genes reported to be hypermethylated in malignant pleural mesothelioma (KAZALD1, TMEM30B and MAPK13). Results: TMEM30B and MAPK13 were not methylated in either case. However, KAZALD1 was highly methylated in sarcomatoid type peritoneal mesothelioma. Discussion: We first report that the KAZALD1 gene was hypermethylated in sarcomatoid type malignant peritoneal mesothelioma.

Citation Format: Ritsuko Oikawa, Yoshiyuki Watanabe, Ryota Hama, Hiroyuki Yamamoto, Fumio Itoh. Characterization of DNA hypermethylation in two cases of peritoneal mesothelioma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1067. doi:10.1158/1538-7445.AM2015-1067
Abstract 1069: DNA methylation in circulating free DNA as a new biomarker for pancreatic cancer
Authors: Shinjo Keiko , Ohka Fumiharu , Katsushima Keisuke , Hatanaka Akira , Ichimura Norihisa , Juan Zhao , Yamao Kenji , Kondo Yutaka
Published: August 2015

Despite the clinical advancement, pancreatic cancer remains one of the most deadly cancers. Pancreatic cancers are often difficult to diagnose and there is no reliable screening biomarker for early detection of this cancer. Epigenetic alterations have emerged as a common hallmark of many types of human cancers as well as pancreatic cancers. Studies have shown that epigenetic dysregulation is associated with multiple steps during carcinogenesis. Therefore, detection of aberrant DNA methylation from blood samples may be a promising diagnostic tool for pancreatic cancers.

First, we analyzed KRAS mutation status of 100 samples obtained by Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA), which is a safe and accurate technique to perform the pancreatic tissue biopsy, and found that more than 80% of the cells in the samples harbors KRAS mutation indicating high amount of cancer cells in the specimens. Next, we performed genome-wide DNA methylation analysis of 38 EUS-FNA specimens using Illumina Infinium HumanMethylation450 BeadChip. We further analyzed the deposited DNA methylation profiles of other tumor types in the public databases, such as TCGA, and identified 8 marker genes with high frequency and specificity of DNA methylation in pancreatic cancers. We validated the DNA methylation status of these genes by bisulfite pyrosequencing and quantitative methylation specific PCR. These markers were frequently methylated in EUS-FNA samples (95% of samples are methylation positive in at least one marker gene). Now we are analyzing the DNA methylation status in pancreatic cancers and corresponding blood samples in order to identify the most effective set of markers for diagnose pancreatic cancers. Our results suggest that analysis of DNA methylation in the circulating free DNA might be a safe and potent tool for evaluating and staging pancreatic cancers.

Citation Format: Keiko Shinjo, Fumiharu Ohka, Keisuke Katsushima, Akira Hatanaka, Norihisa Ichimura, Zhao Juan, Kenji Yamao, Yutaka Kondo. DNA methylation in circulating free DNA as a new biomarker for pancreatic cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1069. doi:10.1158/1538-7445.AM2015-1069
Abstract 1073: KRAS-mediated epigenetic reprogramming upregulates RELB-dependent CXCR2 expression to promote transformation
Authors: Durand Joel , Baldwin Albert
Published: August 2015

In addition to genetic aberrations, cancer cells harbor global epigenetic abnormalities. Mutant KRAS expression has been shown to induce aberrant DNA methylation and silencing of key tumor suppressors. However, nearly all attempts to directly target oncogenic KRAS have been unsuccessful, yielding an impetus for alternative strategies. Furthermore, in the majority of KRAS-driven tumors, the NF-κB pathway is constitutively active and contributes to cellular transformation. Previously, our lab has shown that knocking out the p65 subunit of NF-κB significantly reduces tumor development in a mouse model of KRAS-induced lung cancer. Here we aim to better understand KRAS-mediated epigenetic reprogramming and uncover novel effects on the NF-κB pathway. Our data suggests that KRAS-mediated epigenetic reprogramming alters the NF-κB pathway to promote transformation. Using the Infinium DNA Methylation Assay, we uncovered genome-wide changes in lung and pancreatic cells expressing KRAS(G12V) compared to wild-type cells. Expression of oncogenic KRAS was accompanied by promoter hypermethylation of tumor suppressors (e.g. FAS, BRCA1, IRAK3) and hypomethylation of oncogenes. Interestingly, we observed hypomethylation at the promoter of the RELB subunit of NF-κB in cells expressing KRAS(G12V). Hypomethylation at the RELB promoter correlated with increased expression of CXCR2, an NF-κB target gene. RNAi against RELB and CXCR2 significantly decreased colony growth in lung and pancreatic cancer cell lines expressing mutant KRAS. Together, these results support a role for KRAS in epigenetic reprogramming and suggest that RELB-dependent CXCR2 expression may be critical in KRAS-driven cancers. RELB and CXCR2 represent potential therapeutic targets that could lead to more effective treatments in the context of KRAS-driven malignancies.

Citation Format: Joel Durand, Albert Baldwin. KRAS-mediated epigenetic reprogramming upregulates RELB-dependent CXCR2 expression to promote transformation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1073. doi:10.1158/1538-7445.AM2015-1073
Abstract 1094: Deciphering driver mechanisms for tumorigenesis in BRAF/NRAS double wild-type melanoma through integration of heterogeneous genome-wide datasets
Authors: Mandal Amit , Girotti Maria Romina , Viros Amaya , Gremel Gabriela , Galvani Elena , Lee Rebecca , Lim Kok Haw Jonathan , Furney Simon J. , Lorigan Paul , Marais Richard
Published: August 2015

In cutaneous melanoma, the most lethal form of skin cancer, BRAF is mutated in ∼45% of cases and NRAS in a further ∼20% of cases. The lack of known drivers in the remaining proportion of samples represents a challenge for personalized medicine. We hypothesize that the BRAF/NRAS double wild-type (WT) samples are a heterogeneous group driven by multiple events that arise independently. We performed whole exome sequencing (WES) and complimentary RNA-seq of 23 tumor tissue samples from 21 patients. Six of these tumors were WT for BRAF and NRAS, so we studied these cases further. The SNV load in the WT samples is more variable than the BRAF or NRAS driven melanomas. The median number of SNVs/Mb (± SD) for BRAF mutant (n = 9), NRAS mutant (n = 7) and BRAF WT/ NRAS WT (n = 6) samples were: 12.2 ± 8.0, 42.2 ± 82.7 and 12.6 ± 90.2 respectively. Next we analyzed the TCGA cutaneous melanoma cohort where there are 154 BRAF and 92 NRAS mutated and 76 BRAF WT/NRAS WT samples. The median numbers of SNVs (± SD) were 374 ± 638.3, 577 ± 518.3 and 146.5 ± 1221.6 respectively. The p-value for the comparisons BRAF vs. WT and NRAS vs. WT were 0.087 and 0.002 respectively (two-tailed Mann-Whitney test). We identified the somatic mutations that are specifically enriched for double wild-type samples and observed that the top two hits, NF1 (30.2% in double WT vs. 5.7% otherwise) and KIT (15.8% in double WT vs. 0.8% otherwise) are known driver gene candidates for wild-type melanomas, but we also find other novel candidate driver genes. Thus, we present a framework for identification of driver mutations and therapeutic targets in double wild-type melanomas and integration of these types of data with other large datasets such as those derived from RNA-seq and RPPA will assist in the development of approaches to stratify double wild-type patients for targeted or immune-therapies.

Citation Format: Amit Mandal, Maria Romina Girotti, Amaya Viros, Gabriela Gremel, Elena Galvani, Rebecca Lee, Kok Haw Jonathan Lim, Simon J. Furney, Paul Lorigan, Richard Marais. Deciphering driver mechanisms for tumorigenesis in BRAF/NRAS double wild-type melanoma through integration of heterogeneous genome-wide datasets. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1094. doi:10.1158/1538-7445.AM2015-1094
Abstract 1095: In silico mapping of oncogene networks implicate the WNT pathway in the glioblastoma MES subtype
Authors: Joy Anna M. , Smirnov Ivan , Reiser Mark , Kim Seungchan , Feuerstein Burt
Published: August 2015

It is clear that glioblastoma (GBM) is a collection of molecular subtypes, however little is known of their oncogene signaling. To address this we are currently developing a new method to map PI3K/AKT/mTOR network activity. It uses TCGA reverse-phase protein array data and is based on correlations between active pathway members. This approach provides evidence the WNT pathway is active in the Mesenchymal subgroup and we are currently validating these results. Based on this data we hypothesize Mesenchymal patients will respond to WNT inhibitors.

Citation Format: Anna M. Joy, Ivan Smirnov, Mark Reiser, Seungchan Kim, Burt Feuerstein. In silico mapping of oncogene networks implicate the WNT pathway in the glioblastoma MES subtype. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1095. doi:10.1158/1538-7445.AM2015-1095
Abstract 11: Molecular mechanisms of inverse association between cancer and Alzheimer's disease
Authors: Yang Yoosoo , Lee EunJung , Kim In-San
Published: August 2015

Apoptosis has been implicated in both cancer and neurodegenerative disease such as Alzheimer's disease (AD). Defective or inefficient apoptosis is an acquired hallmark of cancer cells, while cytochrome c-dependent apoptosis is a crucial pathway in AD related neuronal cell death. Notably, recent studies have revealed an inverse correlation between development of cancer and AD—AD patients show a reduced risk of cancer, while cancer survivors are less likely to acquire AD. Here we propose that amyloid β (Aβ), a widely accepted pathological culprit of AD, hijacks the intrinsic apoptotic pathway eliciting different symptoms for cancer and AD. By developing an in vitro single-vesicle cytochrome c release assay, we show that even in the absence of death signal the oligomeric form of synthetic Aβ directly interacts with a pro-apoptotic bcl-2 family protein BAK and induces apoptotic pores sufficient for cytochrome c release. Furthermore, Aβ oligomers not only cause neuronal cell death in HT-22 cells but also the apoptotic effect was significantly reduced with suppressed BAK expression via siRNA, further substantiating its apoptotic interaction with BAK. Moreover, a significant reduction of cytochrome c release was also observed with isolated mitochondria from BAK siRNA-treated HT-22 cells which specifically identifies the target machinery. These results demonstrate Aβ oligomers’ efficacy as a potent apoptotic ligand. In collection, we suggest that while Aβ oligomers may cause neuronal cell death in AD it may also ameliorate tumor proliferation in relation to cancer. Thus, providing molecular level evidence for the inverse association reported between cancer and AD patients.

Citation Format: Yoosoo Yang, EunJung Lee, In-San Kim. Molecular mechanisms of inverse association between cancer and Alzheimer's disease. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 11. doi:10.1158/1538-7445.AM2015-11
Abstract 1105: Clinical associations between altered bladder cancer genes
Authors: Nickerson Michael L. , Im Kate M. , Turan Sevilay , Moore Lee E. , Dean Michael , Theodorescu Dan
Published: August 2015

Next generation sequencing (NGS) has identified frequently altered cancer genes in urologic cancers including bladder cancer (BCa). We examined BCa tumors from 54 U.S. patients using exome NGS and targeted sequencing, and compared these to 99 extensively characterized Chinese patient tumors (Guo et al., 2013). We are first to show that telomerase (TERT) is the gene most frequently altered by somatic sequence changes in 37/54 (69%) of tumors and that TERT is also altered by rare and common germline variants in 30/54 (56%) tumors. Most TERT germline variants were novel (19/20) and three variants were confirmed as both somatic and germline in distinct tumors, including c. - 245 T>C (rs2853669), the most recurrent variant. A subset of variants created or altered transcription factor binding sites and promoter CpGs. Telomere length was significantly shorter in tumor versus adjacent normal tissue from the same patient (p = 0.004) indicating TERT minimally lengthens telomeres in rapidly proliferating BCa cells. Somatic TERT alterations were not associated with stage, grade, or alterations in other BCa genes identified by exome sequencing. STAG2 is altered by somatic mutations in 20/153 (13%) tumors and by X chromosome deletions and promoter hypermethylation. STAG2 mutant tumors had higher levels of aneuploidy (p = 0.01) and Kaplan-Meier analysis indicated a poor prognosis due to significantly shorter survival times (p < 0.001). Somatic BAP1 alterations were associated with papillary histologic features in 15% of Caucasian patient tumors but were infrequently observed in Asian patients (p = 0.003), which remained marginally significant after accounting for differences in tumor grade and stage (p = 0.037). BAP1 mutations co-occurred with histone 3 lysine 27-specific demethylase (KDM6A) mutations (p = 0.017), indicating the cancer genes might provide complementary survival advantages to cancer cells. We find that cancer gene alterations are highly predictive of canonical features of the BCa phenotype and ∼4-5 tumor suppressors are altered for each oncogene. X chromosome cancer genes such as STAG2 and KDM6A have a single copy in men and their lethality (as shown for STAG2) likely contributes to the gender bias in cancer. We speculate that TERT alterations are early events in BCa and identification of somatic alterations in urine may provide a biomarker for early BCa diagnosis. BAP1 mutations contribute to frequent BRCA pathway alterations in BCa and indicate treatment with poly(ADP ribose) polymerase inhibitors would be effective.

Citation Format: Michael L. Nickerson, Kate M. Im, Sevilay Turan, Lee E. Moore, Michael Dean, Dan Theodorescu. Clinical associations between altered bladder cancer genes. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1105. doi:10.1158/1538-7445.AM2015-1105
Abstract 1117: Who's in the driver's seat? Identifying causative variants of colorectal cancer
Authors: Coggins Nicole , Carvajal-Carmona Luis , Segal David
Published: August 2015

Colorectal Cancer (CRC) is one of the leading causes of mortality in the US, being the third most common cancer among both men and women. While an exact cause of CRC has not yet been elucidated, the existence of a clear heritable genetic component has been established. With the recent surge of data from Genome-Wide Association Studies, a handful of CRC-associated loci have been identified via tagSNPs. While the probability of the tagSNPs themselves being the causative variants is low, the key may lie among the number of single-base variants linked to each tagSNP. It is crucial, in order to enhance our understanding CRC tumorigenesis, to extract these functional variants from the pool of associated SNPs and validate their causative significance within the cell.

This project combines bioinformatics and functional experiments to both identify and assess the role of functional variants in CRC pathogenesis. One of the major hurdles in understanding the role of CRC risk-associated SNPs is the fact that virtually all variants lie in Non-Protein Coding Region, often tens of thousands of bases away from flanking genes. By combining variant data from publically available databases, such as that of the ENCODE project, this research presents a method to prioritize all variants within a set of disease-associated loci based on potential regulatory functionality in silico. With this technique, we have identified a handful of SNPs with functional potential that may be functioning specifically in CRC initiation. In parallel, we are in the process of developing a method to effectively produce isogenic cell lines modeling the appropriate variant alleles, utilizing the CRISPR/Cas9 system to stimulate homologous recombination in the presence of a donor template. With this method, transcriptional and regulatory changes within the cell due to the single-base genomic edit can be measured in order to characterize these functional variants. Utilizing this technology, we have been able to create our first SNP model of SNP rs6983267, a known functional variant whose risk SNP promotes CRC development by means of increased binding and activation of WNT signaling pathway elements, c-Myc and β-Catenin. The ultimate goal of this project is to better understand the role of CRC risk-associated variants, the result of which could have substantial clinical relevance in the generation of new therapies and diagnostics for the treatment of CRC.

Citation Format: Nicole Coggins, Luis Carvajal-Carmona, David Segal. Who's in the driver's seat? Identifying causative variants of colorectal cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1117. doi:10.1158/1538-7445.AM2015-1117
Abstract 1122: Personalized oncogenomics in advanced stage breast cancer
Authors: Sun Sophie , Gelmon Karen A. , Chia Stephen , Lohrisch Caroline , Shenkier Tamara , Villa Diego , Shen Yaoqing , Jones Martin , Pleasance Erin , Kasaian Katayoon , Eirew Peter , Leelakumari Sreeja , Ma Yusanne , Ng Tony , Yip Stephen , Jones Steven JM , Marra Marco A. , Laskin Janessa J.
Published: August 2015

Background

Breast cancer is a complex disease with clinical, pathological, and molecular heterogeneity. Recent studies have identified several subtypes of breast cancers driven by specific molecular pathways that can be inhibited by targeted drugs. We studied the feasibility of using molecular data from whole genome and transcriptome sequencing of breast cancers to guide treatment.

Methods

Patients were consented as part of the Personalized Onco-Genomics (POG) study at the British Columbia Cancer Agency. Fresh tumor, blood for normal DNA, and archival tumor were collected. Tissue and blood samples were sequenced using the Ion Torrent AmpliSeq panel, followed by comprehensive DNA and RNA sequencing. In-depth bioinformatic analyses were performed. Somatic mutations, copy number changes, structural variants and gene expression were characterized. Results from genomic analyses were reviewed in a multi-disciplinary team.

Results

From August 2012 to October 2014, tissue samples from 30 patients with advanced stage breast cancer were analyzed. The median age at diagnosis was 53 years (range 32-75). The median number of lines of cytotoxic therapy prior to sequencing was 2 (range 0-10). All cases were invasive ductal carcinoma; 63% were ER+ and HER2-; 27% triple negative; 7% ER+ and HER2+; and 3% ER- and HER2+. The most frequently mutated genes were TP53 (67%), PI3KCA (23%), ESR1 (20%), ATM (10%), ARID1A (10%), and BRCA1/2 (10%). Somatic mutations in other genes of interest including HER2, PARP1, NF1, BAP1, PTEN, NOTCH1, were also identified. Molecular data were informative for patient care and/or actionable to guide treatment in 57% (17/30) of cases.

Conclusion

The use of whole genome sequencing technology to identify valuable molecular information to guide personalized breast cancer treatment is feasible. Further studies are warranted to evaluate the usefulness of genome-wide sequencing of breast cancers in clinical practice.

Citation Format: Sophie Sun, Karen A. Gelmon, Stephen Chia, Caroline Lohrisch, Tamara Shenkier, Diego Villa, Yaoqing Shen, Martin Jones, Erin Pleasance, Katayoon Kasaian, Peter Eirew, Sreeja Leelakumari, Yusanne Ma, Tony Ng, Stephen Yip, Steven JM Jones, Marco A. Marra, Janessa J. Laskin. Personalized oncogenomics in advanced stage breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1122. doi:10.1158/1538-7445.AM2015-1122
Abstract 1126: Differential effects of metformin and phenformin vs. other complex 1 inhibitors in vitro and in vivo
Authors: Algire Carolyn , Ehrmann Alexander , Christian Sven , Neuhaus Roland , Menz Stephan , Schwede Wolfgang , Haerter Michael , Haegebarth Andrea
Published: August 2015

Biguanides, such as metformin and phenformin, are currently under investigation for their potential use as anti-neoplastic therapy. Recent publications suggest that both metformin and phenformin exert effects through inhibition of Complex 1 in the electron transport chain. We investigated the effects of metformin and phenformin compared to rotenone, in vitro, and known Complex 1 inhibitor BAY 872243, in vitro and in vivo.

As expected, rotenone and BAY 872243 showed strong inhibition of Complex I in cell-based and enzymatic assays with IC50 values in the nanomolar range. The high affinity binding to Complex I was also reflected by induction of cellular reactive oxygen species (ROS) with EC50 values in the nanomolar range. In contrast, the biguanides neither inhibited Complex I in cell-based and biochemical assays, nor led to an induction of ROS at concentrations up to 300 μM.

In vivo exposure analysis shows that at the maximal tolerated dose (100 mg/kg QD i.p for phenformin and 350 mg/kg i.p QD for metformin), neither metformin nor phenformin had plasma exposure levels over the IC50 for proliferation in vitro. Recent reports have suggested that biguanides, via the inhibition of Complex 1 and subsequent reduction in oxygen consumption, can be used to re-oxygenate tumor areas prior to radiation therapy. Pimonidazole staining demonstrated that metformin and phenformin effectively eliminated hypoxic regions in NCI-H460 xenografts in a time course dependent manner that reflected the exposure. In contrast to the biguanides, BAY 87-2243 effectively eliminated hypoxic regions up to 24 hours post compound administration. Finally, both phenformin and metformin had minimal effects on inhibition of tumor growth, even in LKB1-deleted xenografts which have been reported to be especially sensitive to biguanides. In conclusion, our in vitro experiments on the mode of action of biguanides raise questions as whether the in vivo effects on hypoxic tumor regions are related to direct inhibition of Complex I.

Citation Format: Carolyn Algire, Alexander Ehrmann, Sven Christian, Roland Neuhaus, Stephan Menz, Wolfgang Schwede, Michael Haerter, Andrea Haegebarth. Differential effects of metformin and phenformin vs. other complex 1 inhibitors in vitro and in vivo. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1126. doi:10.1158/1538-7445.AM2015-1126
Abstract 1135: Loss of HSulf-1 promotes defective autophagy and increased lipid droplet biogenesis in ovarian cancer
Authors: Roy Debarshi , Mondal Susmita , Khurana Ashwani , He Xiaoping , Hammond Edward , Dredge Keith , Shridhar Viji
Published: August 2015

Dysregulation of autophagy and altered metabolic pathways are frequently observed in cancer. Due to these alterations, pharmacological targeting of these two pathways simultaneously could provide a viable therapeutic option. Although the association between these two pathways is well characterized in metabolic disorders, it is not well defined in ovarian cancer (OVCA). In this regard, we found that loss of endosulfatase HSulf-1, a known putative tumor suppressor, suppresses LC3-GFP foci formation and promotes increased lipid droplet (LD) biogenesis suggesting that absence of HSulf-1 in OVCA affects both autophagy and lipid metabolism. While isogenic cells with genetic ablation of HSulf-1 (OV202Sh1/2 and TOV2223Sh1 cells) displayed LDs, the nontargeted control transduced (NTC) OV202 and TOV2223 cells had significantly less LDs. In contrast, Transmission Electron Micrographs (TEMs) showed that OV202 and TOV2223 NTC cells had significantly more autophagic vacuoles (AVs) compared to their isogenic ShRNA targeted cells. Conversely, ectopic expression of HSulf-1 in SKOV3 cells decreased the number of LDs and increased the number of AVs compared to vector transfected controls. Here we report that OV202Sh1 cells and HSulf-1 deficient OV2008 cells have increased p-cPLA2α(ser505) levels that are associated with biogenesis of large number of LDs with reduced AVs. Interestingly, pharmacological inhibition of cPLA2α with AACOCF3 in OV202Sh1 cells resulted in reduced LD biogenesis, inhibited colony formation and reduced tumorigenesis in vivo. More importantly, treatment of HSulf-1 deficient cells with HS mimetic PG545 which can compensate for loss of HSulf-1, reduced LD biogenesis, promoted autophagy and inhibited tumor growth in vivo. Collectively, these results show a critical role of HSulf-1 in regulating both autophagy and LD biogenesis in ovarian cancer.

Citation Format: Debarshi Roy, Susmita Mondal, Ashwani Khurana, Xiaoping He, Edward Hammond, Keith Dredge, Viji Shridhar. Loss of HSulf-1 promotes defective autophagy and increased lipid droplet biogenesis in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1135. doi:10.1158/1538-7445.AM2015-1135
Abstract 1137: Mitochondrial targeting of EGFR-VIII in temozolomide resistant glioblastoma models
Authors: Velpula Kiran Kumar , Asuthkar Swapna , Martin Sarah E. , Lathia Justin D. , Tsung Andrew J.
Published: August 2015

Epidermal growth factor receptor (EGFR) over-expression is the most frequent genetic alteration associated with glioblastoma and has emerged as a highly attractive therapeutic target based on its high frequency of gene amplification and mutation as well as its deregulated cell signaling cascades that drive GBM pathophysiology. The genetic aberration of wtEGFR is often accompanied by the overexpression of a mutant EGFR known as EGFR variant III (EGFRvIII), which is expressed in 30% of GBM tumors. Despite recent innovations, may it be neurosurgical techniques, chemotherapeutics or molecular targeted therapies only few treatments showed promise. Temozolomide (TMZ) is one such and has shown improved survival in GBM when administered with concomitant radiotherapy. However, acquired TMZ chemoresistance occurs in more than 90% of recurrent GBMs. To delineate the mechanisms in the context of TMZ resistance, we generated an aggressive TMZ resistant model by continual exposure of U373 cells constitutively expressing EGFRvIII (U373vIII) to 150uM TMZ for 6 months (U373vIIIR). Our preliminary data showed that dichloroacetic acid (DCA) induced apoptosis and demonstrated reduced cell survival in the context of TMZ resistance. DCA treatment also demonstrated reduced EGFRvIII mitochondrial translocation in U373vIIIR cells. Micro array studies conducted on U373vIII or U373vIIR and their subsequent treatment with DCA revealed that PDK1 is the sole target in TMZ resistant tumors expressing EGFRvIII. Since EGFRvIII expression displays an aggressive phenotype yet remains tumor specific, we propose studying DCA alone or in combination with dasatinib, an inhibitor of EGFR subcellular translocation, in U373vIII and U373vIII-TMZ-R cell lines. By more accurately depicting the status of GBM at the time of recurrence after standard of care chemotherapy treatment, the investigation of DCA will augment and possibly redefine our current understanding of resistance mechanisms and establish novel methods of circumventing this pervasive phenomena.

Note: This abstract was not presented at the meeting.

Citation Format: Kiran Kumar Velpula, Swapna Asuthkar, Sarah E. Martin, Justin D. Lathia, Andrew J. Tsung. Mitochondrial targeting of EGFR-VIII in temozolomide resistant glioblastoma models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1137. doi:10.1158/1538-7445.AM2015-1137