bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1476: Landscape of fusion RNAs generated by cis-spliced adjacent genes in cancer and normal physiology
Authors: Qin Fujun
Published: August 2018

The fusion RNA, SLC45A3-ELK4, was found to be a product of cis-splicing between the two adjacent genes (cis-SAGe). Previously, we used LNCaP, a prostate cancer cell line as a model, and identified 16 additional cis-SAGe events by silencing transcription factor CTCF and paired-end RNA sequencing. One out of the 16 fusion RNAs, D2HGDH-GAL3ST2, is more frequently seen in cancer samples, and seems to be enriched in the African American group. It is more frequently detected in late stage cancer, suggesting a role in cancer progression.

By studying the features associated with the 16 fusions, we developed a set of rules: 1) the parental genes are same-strand-neighboring genes, 2) the distance of between the genes is within 30kb; 3) the 5' genes are actively transcribing; and 4) the fusion RNAs tend to have the second-to-last exon in the 5' genes joined to the second exon in the 3' genes. To investigate the cis-SAGe fusion RNAs at global, we first randomly selected 20 neighboring genes in the genome, and detected four fusion events using these rules in prostate cancer and non-cancerous cells.

To further understand the cis-SAGe fusions in human tissues and cells, we then performed, curated and analyzed nearly 300 RNA-seq libraries covering 30 different non-neoplastic human tissues and cells. More than 10,000 fusion transcripts were found. We focused on the neighboring fusions and performed analysis. Appling our cis-SAGe rules, we identified around 1486 cis-SAGe fusion candidates. A few sequence motifs were found enriched close to the fusion junction sites. We performed functional experiments on a few widely expressed fusions, such as ADCK4-NUMBL, CLN6-CAML4, DUS4L-BCAP29, and found that silencing them resulted in dramatic reduction in moral cell growth and /or motility.

Citation Format: Fujun Qin. Landscape of fusion RNAs generated by cis-spliced adjacent genes in cancer and normal physiology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1476.
Abstract 1480: Overexpression of histone H3.1 induced cell malignant transformation and mutation of H3.1 C96/110A further induced chromatin instability
Authors: Zhu Yusha , Sun Hong , Costa Max
Published: August 2018

Histones are major proteins in chromatin and are important for chromatin structure and gene expression. Canonical histones are defined as a group of replication-dependent histone proteins and are mainly synthesized during S phase of the cell cycle. Their mRNAs are devoid of poly-adenylated tails, but end up with a stem loop structure, which can be targeted by proteins such as stem-loop binding proteins (SLBP) for processing, exportation, and degradation. In our previous studies, arsenic exposure was found to be associated with reduced levels of SLBP and accumulation of poly-adenylated mRNA of canonical histone H3.1, and ultimately led to malignant transformation of cells. Histone H3.1 and H3.2 are canonical histones and they differ by only one amino acid, Ser96 on H3.2 compared with Cys96 on H3.1, and both of them have a Cys110. Histone H3 is the only histone that contains cysteine. Cysteine (C) is a unique amino acid that is rarely used by nature and it has a reactive thiol side chain that can easily form dimers containing disulfide bridge when oxidized. Here it is hypothesized that cysteine is vital for the structure and stability of chromatin. To test this hypothesis, we generated an H3.1 mutant H3.1 C96/110A, where two cysteine residues were replaced by alanine (A). The wild-type H3.1 and H3.1 C96/110A were exogenously expressed in the Beas-2B cells. Interestingly, Beas-2B cells ectopically expressing poly-adenylated H3.1 and H3.1 C96/110A showed decreased proliferation rate with cell cycle arrest at S phase, and cells also acquired cancer cell properties such as increased anchorage-independent growth along with migration and invasion ability. It was confirmed that H3.1 C96/110A could be incorporated into chromatin, resulting in increased chromatin instability and DNA damage. In addition, the RNA-seq assay revealed a total of 678 genes that were oppositely expressed between H3.1 and H3.1 C96/110A ectopically expressed cells. These findings emphasize the importance of a balance among histone variants in the cell and an essential role of cysteines in histone H3.1.

Citation Format: Yusha Zhu, Hong Sun, Max Costa. Overexpression of histone H3.1 induced cell malignant transformation and mutation of H3.1 C96/110A further induced chromatin instability [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1480.
Abstract 1487: Investigation of triple-negative breast cancer tumor conversion through high-content screening approaches
Authors: Gaudeau Albane , Lamamy Véronique , Jaskowiak Anne-Laure , Scerri Xavier , Chanrion Benjamin , Dorval Thierry , Nery Elaine Del , Perez Franck , Camonis Jacques , Stéphan Jean-Philippe
Published: August 2018

Among Breast Cancer (BC) classification, Triple-Negative (TN) form is the most aggressive, lacking efficient and specific treatment. Our project aims at investigating the possibility to convert TNBC tumor cell lines in Estrogen Receptor (ER), Progesterone Receptor (PR) and/or Human Epidermal growth factor Receptor 2 (HER2)-positive cells using various modalities (siRNA, CRISPR-Cas9, small molecules). Our approach consists in optimizing a multi-parametric High-Content Screening (HCS) relying on phenotypic read-outs such as expression of TNBC positive and negative markers, cell behavior and viability. This poster will present our initial results, validation and future plans. This thesis project fits within a context of drug discovery, innovation, comparison of technologies and understanding of key biological processes and pathways, carried out in partnership between Servier Research Institute and Institut Curie.

Citation Format: Albane Gaudeau, Véronique Lamamy, Anne-Laure Jaskowiak, Xavier Scerri, Benjamin Chanrion, Thierry Dorval, Elaine Del Nery, Franck Perez, Jacques Camonis, Jean-Philippe Stéphan. Investigation of triple-negative breast cancer tumor conversion through high-content screening approaches [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1487.
Abstract 1489: Differential gene expression profiles between normal cervical tissue and high-grade squamous intraepithelial lesion expressing p16 and high-risk HPV E6/E7 mRNA
Authors: Benedetti Ines , Barrios Lia , Reyes Niradiz
Published: August 2018

Background:

HPV is necessary but not sufficient for development of uterine cervix carcinoma. There are genetic factors that influence the progression of premalignant lesions. Gene expression profile differences between healthy and premalignant lesion could contribute to identify potential biomarkers for each step of cervical tumor progression.

Design:

A female patient with an initial diagnosis of low grade squamous intraepithelial lesion (SIL) was invited to participate in the study. During follow-up a high grade SIL was observed by colposcopy, and two samples were obtained from the lesion: one sample was processed for histopathologic diagnosis, and additionally stained by IHC with p16 antibody; the other sample, along with a matched sample of healthy tissue taken distantly from the lesion were stored in a nucleic acids preservation reagent for subsequent analysis of gene expression. Differences in gene expression between healthy tissue and high grade SIL tissue were assessed in triplicate using the cDNA gene expression Illumina microarray Human-HT-12-V4. A statistical p value was computed comparing expression levels from the tissue lesion and healthy tissue. Genes with a fold-change ≥ 2 and a p value <0.05 were regarded as differentially expressed. Additionally, two cervix swabs were obtained for detection of HPV DNA by PCR, and for assessment of high risk HPV E6/E7 mRNA expression by RT-PCR.

Results:

Histopathological evaluation of the lesion confirmed a high grade squamous intraepithelial lesion, with positive p16 staining. HPV-18 DNA was detected and E6/E7 HPV 18 mRNA expression was confirmed. Gene expression analysis detected 33 overexpressed genes, including TMEM45A, SPRR2B, KRT16 and RBP1, and 30 underexpressed genes, including SPARCL1, GCNT3, TMPRSS11b

and ESR1, in high grade SIL compared to healthy uterine cervical tissue. Overexpressed genes belonged to important cellular pathways, such as queratinization, defensins, cytoskeletal signaling, cellular adhesion and extracellular matrix remodeling. Underexpressed genes belonged to metabolic pathways, cell trafficking, and chemokine signaling, among others.

Conclusions:

Microarray analysis identified differential expression between healthy cervical tissue and high grade SIL with confirmed HPV-18 transforming infection. Differentially expressed genes belonged to important cellular pathways. Further studies are required to evaluate the usefulness of these genes as risk markers for progression.

Citation Format: Ines Benedetti, Lia Barrios, Niradiz Reyes. Differential gene expression profiles between normal cervical tissue and high-grade squamous intraepithelial lesion expressing p16 and high-risk HPV E6/E7 mRNA [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1489.
Abstract 1496: Different effects of lncRNA NDRG1-OT1_v4 fragments on regulating NDRG1 transcription in breast cancer cells under hypoxia
Authors: Yeh Chingching , Tsai Mong-Hsun , Chuang Eric , Lai Liang-Chuan
Published: August 2018

Hypoxia is a crucial factor in aggressiveness of solid tumor by driving multiple signaling pathways. Recent researchers indicated that long non-coding RNA (lncRNA) could promote or inhibit tumor aggressiveness by regulating gene expression. Previous studies in our laboratory found the expression of lncRNA NDRG1-OT1_v4 significantly increased under hypoxia by next-generation sequencing (NGS). Moreover, it was discovered that the NDRG1-OT1_v4 inhibited NDRG1 at both mRNA and protein levels of NDRG1. At the protein level, NDRG1-OT1_v4 improved NDRG1 degradation via ubiquitin-mediated proteolysis pathway. However, the repressive mechanism of NDRG1 at RNA level was still unknown. In this studies, we found that NDRG1-OT1_v4 decreased the NDRG1 promoter activities when we overexpressed NDRG1-OT1_v4 under hypoxia. The fragment (150-263 nt) of NDRG1-OT1_v4 repressed NDRG1 promoter activity significantly by increasing the binding affinity of hnRNPA1. On the other hand, another fragment (264-392 nt) of NDRG1-OT1_v4 improved NDRG1 promoter activity by recruiting HIF-1 alpha. In conclusion, we found a novel mechanism that different fragments of same lncRNA could cause opposite effects on the identical target gene. Keywords: Hypoxia, LncRNA, NDRG1-OT1_v4, NDRG1, HIF-1 alpha, hnRNPA1

Citation Format: Chingching Yeh, Mong-Hsun Tsai, Eric Chuang, Liang-Chuan Lai. Different effects of lncRNA NDRG1-OT1_v4 fragments on regulating NDRG1 transcription in breast cancer cells under hypoxia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1496.
Abstract 15: PA28γ contributes to tumor angiogenesis through regulation of IL-6 and CCL2 in oral squamous cell carcinoma
Authors: Zeng Xin , Li Jing , Liu Jiajia , Ji Ning , Feng Hui , Chen Qianming
Published: August 2018

Background: PA28γ has been shown to promote tumor development and progression. Although PA28γ has been suggested to play a role in tumor angiogenesis, direct evidence for the molecular mechanisms underlying this phenomenon have not been investigated.

Methods: Oral squamous cell carcinoma (OSCC) stable cell lines with varying levels of PA28γ expression were constructed. EA.hy926 human endothelial cells were co-cultured with these lines to evaluate the influence of PA28γ on EA.hy926 cell proliferation, migration, invasion, and tube formation. Using a subcutaneous xenotransplantation tumor model of under-expressing PA28γ HSC-3 cells, we explored effects on tumor angiogenesis in nude mice. We performed screens of the different angiogenesis-related genes and cytokines being expressed using PCR and cytokine chips to identify candidates with the potential to affect tumor angiogenesis. Then, candidates were verified using Western blot, and neutralization tests immunohistochemistry. Finally human OSCC clinical cohort was used to validate.

Results: PA28γ could enhance the ability of OSCC cells to interact with endothelial cells, thereby recruit endothelial cells and induce angiogenesis. Ablation of PA28γ inhibits tumor-induced angiogenesis in xenografts mice model in vivo. In vivo zebrafish model experiments showed that depleted PA28γ remarkably blocked the formation of intersegmental vessels in zebrafish embryos. Furthermore, Angiogenesis-related PCR, cytokine chips and Western blot analysis indicated that PA28γ through regulate IL-6 and CCL2 expression, thereby subsequent activation of the Stat3/VEGF and PI3K/MMP-1 axis to promote angiogenesis. Neutralization or block results IL-6 and CCL2 has an essential role in PA28γ-controlled promotes of angiogenesis. In addition, clinical OSCC cohort validate that PA28γ positively correlates with IL-6 or CCL2 expression.

Conclusion: Collectively, our data show that PA28γ contributes to the tumor angiogenesis, by regulating IL-6 and CCL2. PA28γ thus represents a novel therapeutic target for treating PA28γ-positive OSCC cancers.

Citation Format: Xin Zeng, Jing Li, Jiajia Liu, Ning Ji, Hui Feng, Qianming Chen. PA28γ contributes to tumor angiogenesis through regulation of IL-6 and CCL2 in oral squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 15.
Abstract 150: Elucidating the molecular mechanisms of cutaneous melanoma development from melanocyte stem cells
Authors: Moon Hyeongsun , Donahue Leanne , Zhu Jerry , White Andrew
Published: August 2018

Cutaneous melanoma is the most lethal form among all skin cancers. Multiple genetic etiologies have been identified in primary and metastatic tumors, however, the cellular origin for cutaneous melanoma and their related molecular mechanisms required for early steps of melanoma initiation have not been well defined. Although great progress has been made in the development of targeted therapies for malignant melanoma, knowledge on the factors that can cause tumor formation would provide valuable insight for melanoma prevention and control. Here, our study aims to determine potential means of prevention by identifying the extrinsic environmental factors and cellular conditions responsible for melanoma initiation particularly from tumor-competent melanocyte stem cells. In the study, adult stem cells have been discovered as a compelling candidate for melanoma formation. We also found that quiescent stem cells are relatively unable to develop melanocytic tumors. On the other hand, environmental and physiological stresses can significantly alter the status of melanocyte stem cells which in turn causes melanoma development. Screening the genetic changes that occur in the melanoma-competent stem cells in tumor initiation stages can help to elucidate which molecular pathways may cause the formation of cutaneous melanoma. Our data will provide a new framework for the early initiation steps of cutaneous melanoma from quiescent but tumor-prone stem cells, with the ultimate goal to find better preventative measurements and targeted therapy for cutaneous melanoma.

Citation Format: Hyeongsun Moon, Leanne Donahue, Jerry Zhu, Andrew White. Elucidating the molecular mechanisms of cutaneous melanoma development from melanocyte stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 150.
Abstract 1507: Multiple mechanisms of Myb gene activation revealed through transcriptome analysis of salivary gland adenoid cystic carcinoma
Authors: Frerich Candace , Brayer Kathryn , Painter Brandon , Ness Scott
Published: August 2018

Adenoid cystic carcinoma (ACC) of the salivary gland is a rare head and neck malignancy with poor long-term prognosis. Hallmark chromosomal translocations result in MYB to NFIB (t(6;9)) and MYBL1 to NFIB (t(8;9)) gene fusions. Gene rearrangement and aberrant expression of the MYB and MYBL1 genes, along with the encoded transcription factors, are the oncogenic driver in the majority of ACC tumors. Previous RNA-sequencing (RNA-seq) data collected from FFPE fixed ACC tumors illustrated MYB and MYBL1 elicit similar gene expression profiles in ACC, suggesting these transcription factors drive ACC oncogenes via similar mechanisms. Using these RNA-seq data we have identified EN1 and SOX4 as important Myb regulated genes and demonstrated activation of their promoters in reporter gene assays. We hope to utilize these reporters in high-throughput screens to identify new drug candidates for ACC. We have utilized RNA-seq data to identify an additional mechanism leading to MYB overexpression that appears unique to ACC tumors. In these tumors transcription of the MYB gene is aberrantly activated through use of an alternative promoter located within the first intron of the gene. Transcripts originating from this promoter have been confirmed in an ACC tumor via 5'RACE and the resulting Myb protein has a small N-terminal truncation eliminating the epitope recognized by a common Myb antibody. The alternative promoter is active and regulated in a cell type specific manner in reporter gene assays. Previous studies have shown chromosomal translocations juxtapose NFIB enhancers to activate MYB gene transcription. ChIP and 3C assays in fresh frozen ACC tumors will be utilized to test the hypothesis that NFIB enhancers are specifically recruited to the alternative MYB promoter, thereby activating gene transcription. Thus, multiple mechanisms lead to the aberrant over-expression and truncation of MYB and drives ACC oncogenesis.

Citation Format: Candace Frerich, Kathryn Brayer, Brandon Painter, Scott Ness. Multiple mechanisms of Myb gene activation revealed through transcriptome analysis of salivary gland adenoid cystic carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1507.
Abstract 151: Phenotypic selection screening reveals Annexin A2 as a pancreatic cancer stem cell target
Authors: Hurt Elaine M. , Flynn Matt , Hynes Paul , Cabrera M. Carla , Thomas Suneetha B. , Vlerken-Ysla Lilian van , Slater Louise , Sandercock Alan , Rust Steven , Minter Ralph , Herbst Ronald
Published: August 2018

Cancer stem cells (CSCs) represent an important clinical entity given their role in tumor initiation, metastasis and patient relapse. Phenotypic selection allows for identification of novel antigens important in biological processes. Therefore, we undertook a phenotypic selection campaign using a designed ankyrin-repeat (DARPin) library to identify new cancer stem cell (CSC) targets. CSCs generated from late-stage pancreatic cancer patient-derived xenograft (PDX) models were used to pan a designed Ankyrin repeat (DARPin) library. The most prevalent target identified in this screen was Annexin A2 (ANXA2). ANXA2 is a member of the Annexin family that has diverse roles in cancer including proliferation and invasion and is widely expressed in cancer. Furthermore, expression of ANXA2 is linked to poor prognosis in pancreatic cancer. Targeting of Annexin A2 on the surface of pancreatic cancer cells using DARPins resulted in decreased frequency of CSCs both in vitro and in vivo in several pancreatic patient-derived xenografts. ANXA2 DARPins also reduced proliferation and tumor growth of HPAC cells both in vitro and in vivo. Mechanistically, we found that blocking ANXA2 resulted in decreased IL6 and IL8 secretion, two cytokines that have been linked to CSC expansion and survival. Additionally, we found that pharmacological blockade of ANXA2 resulted in decreased plasmin activation, a pathway that has been linked to dissemination of cancer cells. Together our data suggests that targeting of cell surface ANXA2 is effective at reducing CSCs and could be a novel target for the treatment of pancreatic cancer.

Citation Format: Elaine M. Hurt, Matt Flynn, Paul Hynes, M. Carla Cabrera, Suneetha B. Thomas, Lilian van Vlerken-Ysla, Louise Slater, Alan Sandercock, Steven Rust, Ralph Minter, Ronald Herbst. Phenotypic selection screening reveals Annexin A2 as a pancreatic cancer stem cell target [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 151.
Abstract 1514: Role of polycomb proteins in neuroendocrine prostate cancer
Authors: Connelly Zachary , Yang Shu , Blankenship Anthony , Yu Xiuping
Published: August 2018

Prostate cancer (PCa) is the most common non-skin cancer and the 3rd leading cause of cancer-related death in American men. Androgen ablation therapy has been the gold standard for treating advanced stage PCa since the 1960s. Initially, the PCa patients respond to hormone ablation very positively. However, over time these tumors almost always become resistant to androgen ablation therapy, and tumors begin to grow again. Recent studies found after failure of the new drugs that block androgen action, PCa has increased neuroendocrine phenotype. Understanding the mechanisms through which PCa cells gain neuroendocrine phenotype is critical for the development of novel therapeutics. Polycomb repressive complex are involved in early embryo development and carcinogenesis in several types of cancer including PCa. In this study, we evaluated the involvement of Polycomb proteins in neuroendocrine PCa. Using immunohistochemistry staining, we examined the expression of Polycomb protein EZH2 in advanced stage PCa. We found that EZH2 is highly expressed in both human and mouse neuroendocrine PCa. Also, we found when PCa cells are induced to undergo neuroendocrine differentiation, multiple Polycomb genes are induced. These data suggest that Polycomb genes play a role in the development of neuroendocrine PCa.

Citation Format: Zachary Connelly, Shu Yang, Anthony Blankenship, Xiuping Yu. Role of polycomb proteins in neuroendocrine prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1514.
Abstract 1515: NCOA3 promotes tumor growth by activating telomerase reverse transcriptase and predicts poor prognosis in hepatocellular carcinoma
Authors: Chen Miao , Li Wenbin , Li Yixin , Zhang Changlin , Qin Ge , Long Qian , Shi Dingbo , Guo Wei , Shi Shuihan , Zhang Kefang , Deng Wuguo
Published: August 2018

Hepatocellular carcinoma (HCC) is one of the most common cancers and a leading cause of cancer death worldwide. The rising incidence and mortality rate of HCC in most countries are due to an increased rate of chronic Hepatitis C virus infection, alcohol abuse and obesity. The molecular mechanisms leading to the development of HCC are complicated, consist of various genetic and epigenetic changes, and are associated with altered signaling pathways. Among these molecular events, elevated expression of human telomerase reverse transcriptase (TERT) is an important alteration in HCC and a hallmark of human cancers. TERT is highly expressed in HCC and positively correlated with the development of HCC. However, the detailed mechanism of TERT activation in HCC remains largely unclear. Here, we identified nuclear receptor coactivator-3 (NCOA3) as a TERT promoter-binding protein, which specifically bound to the -234~-144 region of the TERT promoter. Overexpression of NCOA3 enhanced TERT promoter activity and expression, and promoted HCC cell growth. Moreover, we demonstrated that NCOA3 interacted with the transcription factor SP1 to promote TERT expression. The clinical data showed that the expression of NCOA3 and TERT were positively correlated and elevated in HCC tissues, and HCC patients with high level of NCOA3/TERT had poor prognosis. Collectively, our study revealed that NCOA3 bound to the TERT promoter and activated TERT expression to promote HCC growth, indicating that the NCOA3/TERT signaling pathway could be a potential therapeutic target for HCC. [Funding support: This work was supported by the funds from the National Natural Science Foundation of China (81772925, 81472178, 81702761), the State “973 Program” of China (2014CB542005), and the Natural Science Foundation of Guangdong Province (2016A03031100, 2015A030313018, 2017A030313615)].

Citation Format: Miao Chen, Wenbin Li, Yixin Li, Changlin Zhang, Ge Qin, Qian Long, Dingbo Shi, Wei Guo, Shuihan Shi, Kefang Zhang, Wuguo Deng. NCOA3 promotes tumor growth by activating telomerase reverse transcriptase and predicts poor prognosis in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1515.
Abstract 1520: Regulation of stemness by perturbation of OCT4-PP1 axis reduces malignancy of embryonal carcinoma
Authors: Song Bomin , Jang Hyonchol
Published: August 2018

OCT4 (encoded by pou5f1) is a master regulator of pluripotency and closely related to tumor malignancies. Its associations with cancer drug-resistance, recurrence and poor prognosis of the patients has been widely reported. As OCT4 transcriptional activity is directly and indirectly involved in the maintenance of cancer stemness, inhibition of OCT4 activity could be a strategy to reduced cancer malignancies by eliminating stemness. In this study, we investigated a way to regulate the stemness of embryonal carcinoma (EC) by perturbation of posttranslational modification of OCT4. First, to confirm the role of OCT4 in the maintenance of stemness in ECs, we generated the cells which are downregulated in OCT4 doxycycline-dependently. Down-regulation of OCT4 caused weaker alkaline phosphates staining, reduced tumor sphere formation, and reduced expression of stemness marker proteins. Also, the cells underwent retardation of proliferation, cell-cycle arrest in G1 phase, and eventual apoptosis by OCT4 downregulation. The most altered genes in ECs by OCT4 downregulation were networked to signaling pathways regulating pluripotent stem cells, Hippo signaling pathways and transcriptional misregulation in cancer. Second, to regulate OCT4 transcriptional activity, we used phosphorylation of serine 236th (S236) on OCT4. ECs cells, with which their endogenous OCT4 was substituted with an OCT4 phosphorylation-mimicking mutant (Serine 236 to aspartate; S236D), showed similar phenotype to those cells that were depleted in OCT4. Chemical inhibition or knockdown of PP1 (protein phosphatase 1), which remove the phosphorylation, induced accumulation of S236 phosphorylation, and also caused similar phenotype to those cells that were depleted in OCT4. Finally, to evaluate whether regulation of OCT4 can reduce tumor malignancy, we tested whether drug-tolerant persister cancer cells could be reduced by regulation of OCT4. Co-treatment of inhibits PP1 activity with cisplatin significantly reduced drug-resistant colonies compared with single treatment of cisplatin. These results suggested that stemness of ECs could be regulated by OCT4 posttranslational modifications and regulation of cancer stemness could reduce tumor malignancy.

Citation Format: Bomin Song, Hyonchol Jang. Regulation of stemness by perturbation of OCT4-PP1 axis reduces malignancy of embryonal carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1520.
Abstract 1544: Subcutaneous adiposity is an independent predictor in metastases patients
Authors: Tsang Ngan M.
Published: August 2018

Background: The prognostic significance of muscle and adipose tissue in cancer patients provide an appropriate assessment of body composition in recent years. We investigated the independent prognostic significance of visceral and subcutaneous adipose on survival in metastases patients. Methods: We measured visceral adipose tissue index (VATI), subcutaneous adipose tissue index (SATI), and muscle index in 959 metastases patients via computed tomography at L3 level. Univariate and multivariate cox proportional hazards model were used to determine mortality hazard ratio (HR) and were presented in corresponding 95% CI. Result: SATI and gender are independent predictors among patients with metastases disease. High SATI linked to lower mortality (HR:0.749; 95% CI: 0.640-0.877; p<0.001) and longer overall survival (8.38 months; 95% CI, 0.10-137.42) in compared to low SATI patients (4.44 months; 95% CI, 0.03-108.36). Conclusion: The body composition appeared to be true determinants of the prognosis in metastases patients. The research provides an understanding of the association between subcutaneous adipose and the overall survival in metastases patients.

Citation Format: Ngan M. Tsang. Subcutaneous adiposity is an independent predictor in metastases patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1544.
Abstract 1564: Detecting low allele frequency somatic mutations using targeted amplicon sequencing and a novel double strand molecular barcoding scheme
Authors: Zhang Li Jacey , Lee Lucie Siu , Liu Yang Lily , Liu Jeff , Chen Tao , Liu Guoying , Liu Zhitong
Published: August 2018

Using next generation sequencing (NGS) with a noise reduction mechanism to detect somatic variants of 0.1% minor allele frequency or lower has attracted increasing interest for cancer research and diagnostics. Molecular barcoding is a technology that aims to reduce the noise and errors caused by PCR and sequencing artifacts, and improves mutation detection accuracy. Here, we report a novel multiplex PCR-based, ultra-sensitive molecular barcoding technology that drastically reduces false positive calls, additionally, barcoded sequencing reads can be assigned back to both the forward and reverse strands of the original DNA fragment, thus further remove variant call noise. While providing a simple and fast workflow, this technology does not have the base bias associated with capture hybridization. It achieves an error rate of 4 X 10-7 nucleotides, and detects allele frequencies at 0.1%.

Citation Format: Li Jacey Zhang, Lucie Siu Lee, Yang Lily Liu, Jeff Liu, Tao Chen, Guoying Liu, Zhitong Liu. Detecting low allele frequency somatic mutations using targeted amplicon sequencing and a novel double strand molecular barcoding scheme [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1564.
Abstract 1567: Ku70, Ku80 and Rab5C as biomarkers for radiotherapy resistance in rectal tumors during neoadjuvant therapy
Authors: Martins Vilma Regina , Baptistella Antuani Rafael , Silva Petrus Paulo , Cassinela Edson Kuatelela , Marchi Fabio , Leme Adriana Fp , Begnami Maria Dirlei , Aguiar Samuel
Published: August 2018

Rectal cancer is the second most common cancer in the large intestine, corresponding to one-third of the total cases of colorectal cancer. The neoadjuvant chemoradiation in rectal cancer treatment reduces the risk of local recurrence, but pathologic complete response is achieved only in 10-40% of patients. The response to neoadjuvant treatment is poorly understated in rectal tumors as well as the mechanisms associated with chemoradiation resistance. In order to identify the players involved with the therapy resistance, we performed proteomic analysis of biopsies from stage II and III rectal adenocarcinoma patients before neoadjuvant treatment and compared with residual tumor tissue removed by surgery after neoadjuvant therapy. Ku70, Ku80 and Rab5C showed a significant increased expression after the chemoradiation treatment. To better understand the mechanism associated with therapy resistance, we generated an ionizing radiation-resistant rectal adenocarcinoma cell line. These cells and also their secreted exosomes exhibit overexpression of the same three proteins identified in tissue samples from patients submitted to chemoradiation. Using this in vitro model, we identified that Rab5C modulates EGFR internalization promoted by irradiation, which was also essential to regulate the expression of the DNA repair proteins Ku70 and Ku80, and cell resistance to ionizing radiation. Taken together, these findings indicate that EGFR and Rab5C are potential targets to sensitize rectal cancer cells to neoadjuvant treatment. In addition, the presence of Ku70, Ku80 and Rab5C in circulating exosomes can be explored as biomarkers to follow resistance to radiotherapy in patients under neoadjunat therapy for rectal tumors.

Citation Format: Vilma Regina Martins, Antuani Rafael Baptistella, Petrus Paulo Silva, Edson Kuatelela Cassinela, Fabio Marchi, Adriana Fp Leme, Maria Dirlei Begnami, Samuel Aguiar. Ku70, Ku80 and Rab5C as biomarkers for radiotherapy resistance in rectal tumors during neoadjuvant therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1567.
Abstract 158: GSKJ4, a selective jumonji histone H3 lysine27 demethylase inhibitor, targets ovarian cancer stem cells
Authors: Sakaki Hirotsugu
Published: August 2018

Background/Aim: Global increase in the trimethylation of histone H3 at lysine 27 (H3K27me3) has been associated with the differentiation of normal stem cells and cancer cells, however, the role of H3K27me3 in the control of cancer stem cells (CSCs) remains poorly understood. We investigated the impact of increased H3K27me3 on ovarian CSCs using GSKJ4, a selective jumonji H3K27 demethylase inhibitor.

Materials and Methods: The effect of GSKJ4 on the viability as well as on the self-renewal and tumor-initiating capacity of CSCs derived from A2780 and TOV21G human ovarian cancer cell line, was examined.

Results: GSKJ4 induced cell death in A2780 CSC and TOV21G CSC at a concentration non-toxic to normal human fibroblasts. Furthermore, GSKJ4 induced differentiation and inhibited sphere formation as well as stem cell marker expression of A2780 CSC and TOV21G CSC that survived GSKJ4 treatment.

Conclusion: Using ovarian CSCs as a model, we have demonstrated for the first time that GSKJ4 can target CSCs, suggesting a critical role for H3K27 methylation in their maintenance and survival. Our findings thus provide an initial clue to explore the role of GSKJ4 as a potent CSC-targeting agent for ovarian cancer and other types of human cancer.

Citation Format: Hirotsugu Sakaki. GSKJ4, a selective jumonji histone H3 lysine27 demethylase inhibitor, targets ovarian cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 158.
Abstract 1582: The coupling of MDSCs with a computational neural network (NN) to detect solid tumors
Authors: Dominguez George A. , Maslar Kristen , Polo Alexander , Sholevar Cyrus , Campisi Anthony , Roop John , Gabrilovich Dmitry , Rauscher Frank , Kumar Amit
Published: August 2018

The goal of this study was to create a non-invasive cancer detection assay that analyzes flow cytometry data in an objective method. By using an artificial neural network (NN), we were able to distinguish between cancer patients (CPs) and benign/healthy donors (HDs) based upon the flow cytometry profiles of MDSCs and other leukocytes. Myeloid-derived suppressor cells (MDSCs) are known to be key contributors in supporting tumor progression and tumor escape through their ability to suppress anti-tumor responses mediated through T cell and natural killer (NK) cell activity. Several studies have demonstrated their utility as indicators of tumor progression and possible predictors of clinical outcomes, but there is significant overlap with healthy individuals preventing discrete and accurate calls. We used standard multiparametric flow cytometry techniques to immunophenotype MDSCs and other leukocytes found in the peripheral blood of 65 biopsy-confirmed CPs with solid tumors and 84 HDs. A series of NNs utilizing pattern recognition computational algorithms are then created using three data sets: 1) the training set - this ‘teaches' the two output categories of cancer and not cancer, 2) the validation set - this uses backpropagation to improve the accuracy of the trained networks, and 3) the testing set - this is used to rank the trained networks against each other. Finally, a naïve testing set is then used to determine the overall sensitivity and specificity for the top-ranking networks. Using traditional flow cytometry gating methods to analyze MDSCs as a biomarker for cancer detection, it is difficult to achieve high sensitivity and specificity due to the substantial overlap with healthy individuals. Here, we incorporated a standard 12 marker flow cytometry assay with NN technology to achieve a sensitivity of 92% and a specificity of 89%. Pairing the advanced analytical capabilities of our NN with surface biomarker based analysis of MDSCs and certain leukocytes measured in peripheral blood has enabled us the ability to objectively identify patterns indicative for the existence of a solid tumor.

Citation Format: George A. Dominguez, Kristen Maslar, Alexander Polo, Cyrus Sholevar, Anthony Campisi, John Roop, Dmitry Gabrilovich, Frank Rauscher, Amit Kumar. The coupling of MDSCs with a computational neural network (NN) to detect solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1582.
Abstract 1585: Evaluation of circulating free DNA in non-metastatic breast cancer
Authors: Araujo Jhajaira M. , Ponce Jaime , Murillo Alexis , Rebaza Pamela , Danos Pierina , Aguilar Alfredo , Fujita Ricardo , Gomez Henry L. , Pinto Joseph A. , Buleje Jose
Published: August 2018

Background Circulating free DNA (cfDNA) is becoming an important tool in diagnosis and prognosis in several types of cancer. In this study we evaluated the association between cfDNA concentrations in peripheral blood with tumoral features of non-metastatic breast cancer patients.

Methods cfDNA was extracted from plasma of peripheral blood of patients with non-metastatic breast cancer and healthy controls. In patients, blood samples were collected before and after surgery, while in controls samples were collected after their results of breast cancer screening. Detection of copy number of PUM1 and RNaseP genes were performed in a QuantStudio® 3D Digital PCR System and quantification was obtained with QuantStudio® 3D AnalysisSuite™ Cloud Software. Raw values were compared among the different clinicopathologic features.

Results In total 26 patients and 25 controls were included. There were significant differences between controls and patients for levels counts of PUM1 and RNaseP genes (p<0.0001 in both cases). In patients, median concentrations before and after surgery for PUM1 were 10.6 copies/uL (range: 2.2-16.3) vs. 7.6 copies/uL (range: 1.3-12.8), respectively (P=0.04) and for RNaseP were 6.6 copies/uL (range: 0.9-20.0) vs. 5.3 copies/uL (range: 0.5-16.0), respectively (P=0.103). Hormonal tumors produced more cfDNA in terms of RNaseP at diagnosis (7.9 vs. 5.6, P=0.049). When we compared patients according to their hormonal status with controls, RNaseP was able to identified positive hormonal receptors patients (P<0.0001). There were no significant differences between other clinicopathologic features. There was a significant decrease after surgery of RNaseP levels in node negative tumors (P=0.049) and PUM1 levels in >10mm tumors (P=0.041).

Conclusions Our results suggest that PUM1 is a better diagnostic biomarker for non-metastatic breast cancer because RNaseP loss sensitivity in negative hormone receptor cases. Hormonal tumors produced more circulating free DNA. RNaseP could be used to monitor node negative patients and PUM1 to patients with >10mm tumors.

Citation Format: Jhajaira M. Araujo, Jaime Ponce, Alexis Murillo, Pamela Rebaza, Pierina Danos, Alfredo Aguilar, Ricardo Fujita, Henry L. Gomez, Joseph A. Pinto, Jose Buleje. Evaluation of circulating free DNA in non-metastatic breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1585.
Abstract 1599: Capture of tumor cells in blood with "Universal CTC-chip"
Authors: Yoneda Kazue , Kuwata Taiji , Tanaka Fumihiro
Published: August 2018

Background: Circulating tumor cells (CTCs) are not only a surrogate of distant metastasis, but also useful for material of liquid biopsy. "Universal" CTC-chip is a polymeric microfluidic device that can capture CTCs according to their surface marker from whole blood. We have examined the capture efficiency of lung cancer and mesothelioma cells in the blood with CTC-chip coated with antibodies against various surface marker.

Methods: Lung cancer cell lines (PC-9) or mesothelioma cell lines (ACC-MESO-4, MSTO-211H) were used in this study. Cells were CFSE-labeled and added in the blood of healthy donor. A polymeric CTC-chip was coated with two steps: base- and capture-antibody. As capture antibody, we used anti-EpCAM, podoplanin, and EGFR antibody. After flowing these samples, capture efficiency was calculated (number of captured cells/number of flowed cells).

Results: Capture efficiency of PC-9, MESO-4, and METO-211H was 100.0%, 9.5% and 9.1% (EpCAM-chip; anti-EpCAM antibody clone HEA125); 5.8%, 85.9% and 9.0 % (podoplanin-chip; anti-podoplanin antibody clone E1); 38.6, 112.5%, 11.7% (podoplanin-chip; anti-podoplanin antibody clone NZ-1); 30.2%, 21.8%, 38.5% (EGFR-chip; anti-EGFR antibody clone 528), respectively.

Conclusion: Using "Universal CTC-chip" with various antibodies, it was possible to capture nonepithelial tumor cells such as mesothelioma cells.

Citation Format: Kazue Yoneda, Taiji Kuwata, Fumihiro Tanaka. Capture of tumor cells in blood with "Universal CTC-chip" [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1599.
Abstract 1620: ATR inhibition for the treatment of ATM-deficient gastric cancer
Authors: Cai Mu-Yan , Reavis Hunter D. , Chen Jie-Wei , Ganesa Chirag , Kochupurakkal Bose , Shapiro Geoffrey I. , D'Andrea Alan D.
Published: August 2018

ATM and ATR are central kinases in the cellular response to DNA damage and play a crucial role in maintaining genome integrity. Inactivation of ATM has been observed in multiple tumor types and pre-clinical studies suggest that inhibition of ATR is synthetic-lethal in ATM-deficient tumors. More than 10% of gastric cancers carry potentially deleterious mutations in ATM and patients with ATM-deficient gastric cancers could potentially benefit from ATR inhibitor-based therapy. This study focuses on establishing the critical parameters for clinical development of ATR inhibitors in treating gastric cancer. First, we tested the cytotoxic response to ATR inhibition of ATM-deficient and ATM-complemented derivative of human fibroblasts. Next, we utilized a panel of gastric cancer cell lines that represent the spectrum of ATM mutations and ATM expression levels to further confirm the correlation between reduced ATM activity and the cytotoxic effects of ATR inhibition. Third, we evaluated the effect of a series of missense and truncating mutations in ATM that were identified in The Cancer Genome Atlas (TCGA) project. Finally, we developed an immunohistochemistry (IHC) assay to assess the expression level of ATM in gastric tumor samples. Our results confirmed that ATM-deficient human fibroblasts were acutely sensitive to ATR inhibition compared to ATM-complemented cells. Acute sensitivity to ATR inhibition was also observed in gastric cancer cell lines that carry deleterious mutations in ATM and in those that express very low levels of ATM. Complementation experiments with mutant versions of the ATM ORF using ATM-deficient human fibroblasts identified deleterious mutations in ATM that sensitize to ATR inhibition. The IHC assay was applied to a cohort of 208 primary gastric cancers. ATM expression was undetectable in approximately 11% of the samples; an additional 26% expressed very low levels. Our findings suggest that ATR inhibition may elicit favorable responses in gastric cancer patients with tumors harboring deleterious ATM mutations or very low levels of ATM expression. Targeted exon-sequencing methods that are used widely combined with IHC for ATM may be useful for patient selection.

Citation Format: Mu-Yan Cai, Hunter D. Reavis, Jie-Wei Chen, Chirag Ganesa, Bose Kochupurakkal, Geoffrey I. Shapiro, Alan D. D'Andrea. ATR inhibition for the treatment of ATM-deficient gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1620.