bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 2422: Genetic aberrations of Anaplastic Lymphoma Kinase (ALK) are functional drivers in head and neck squamous cell carcinoma (HNSCC)
Authors: Wang Lan , Yeung Thomas Chun Kit , Ngan Hoi-Lam , Poon Peony Hiu Yan , Lui Vivian Wai Yan 
Published: July 2021

Abstract
Head and neck squamous carcinoma (HNSCC) is a cancer with high genetic heterogeneity. Fusions of Anaplastic Lymphoma Kinase (ALK) have been widely described as a functional driver for lung cancer. Yet, mutations and amplification of ALK are found in HNSCC as well as many other human cancers. Their biological impacts on cancer cell growth are underexplored.
Using the Cancer Genome Atlas (TCGA) HNSCC dataset (Firehose Legacy; N=510), we identified a subset of HNSCC patients harboring ALK aberrations (4%), including 0.4% cases with gene amplification and 3.5% with somatic mutations. Transcriptome analysis showed significant tumor-specific ALK upregulation in 43 adjacent normal-HNSCC tumor pairs (2-fold, p=0.001***, TCGA HNSCC Firehose Legacy). Interestingly, clinical analysis revealed that ~94% HNSCC patients with ALK amplification and mutations have advanced diseases (stage III or IV). Among the HNSCC-associated ALK aberrations, ALK amplification, as well as ALK p.R401L mutations are recurrent events. Thus, we sought to investigate their biological functions by ectopic expressions into an HNSCC cell line, FaDu, via retroviral infection. We found that overexpression of ALK WT gene (mimicking gene amplification) and ALK p.R401L significantly promoted cell proliferation vs. EGFP-control (p=0.0004***, 0.01*, respectively). Clonogenic assay also demonstrated both events to be drivers for colony survival vs. EGFP-control by 120%, 150% (p=0.02*, 0.0025**, respectively). Lastly, both events markedly increase FaDu cells invasiveness by Matrigel invasion assay by 67%, 58% (vs. EGFP-control, p&lt;0.0001****, 0.0001****, respectively). In conclusion, ALK aberrations are functional drivers promoting HNSCC aggressiveness.
Acknowledgements: This study is funded by General Research Fund, Research Grant Council, Hong Kong Government, Hong Kong SAR (#17121616 ). VWYL also receives funding supports from General Research Fund, Research Grant Council, Hong Kong Government, Hong Kong SAR (#17121616, #14168517), Research Impact Fund (#R4015-19F, R4017-18), the Health and Medical Research Fund (HMRF#15160691) from the Health and Medical Research Fund, the Food and Health Bureau, the Government of the Hong Kong Special Administrative Region), University-Industry Collaboration Programme (UIM/329; Innovation and Technology Fund, Hong Kong Government, Hong Kong SAR), and the Hong Kong Cancer Fund, Hong Kong SAR.
Citation Format: Lan Wang, Thomas Chun Kit Yeung, Hoi-Lam Ngan, Peony Hiu Yan Poon, Vivian Wai Yan Lui. Genetic aberrations of Anaplastic Lymphoma Kinase (ALK) are functional drivers in head and neck squamous cell carcinoma (HNSCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2422.
Abstract 2427: Identification of transcription factors activated in glioblastoma multiforme
Authors: Wu Zexun , Schreiner Shannon , Farnham Peggy J. , Rhie Suhn K. 
Published: July 2021

Abstract
Introduction Glioblastoma multiforme (GBM) is the most common intrinsic brain tumor in adults and is universally fatal. Our strategy to study the molecular mechanisms leading to GBM is to investigate the role of transcription factors (TFs) that are differentially expressed between normal and GBM. Previous studies have shown that ZFX is a key TF that shows increased expression in GBM. However, the role of ZFX, and the identity of TFs that may work with ZFX, in driving GBM carcinogenesis is not yet understood.
Materials and methods: ChIP-seq was performed to determine ZFX binding sites in the U87 GBM cancer cell line. RNA-seq datasets generated from GBM tissues by The Cancer Genome Atlas (TCGA) were analyzed to identify TFs showing increased expression in GBM. Publicly available H3K27ac chromatin immunoprecipitation (ChIP-seq) and Assay for Transposase Accessible Chromatin with high-throughput sequencing (ATAC-seq) datasets generated from GBM tissues were analyzed to identify active enhancers and TFs that bind to GBM enhancers.
Results: We found that a majority of ZFX binding sites in GBM are located at promoters, with ~1000 binding sites being at non-promoter regions. An overlap analysis of ZFX binding sites in multiple cancer cell types showed that a high percentage of ZFX-binding sites at promoter regions are shared across cell types whereas hundreds of GBM-specific binding sites are located at non-promoter regions. Using TCGA GBM RNA-seq datasets, we next identified differentially expressed TFs. For example, we found that MTY1L and NEUROD2, which have been reported to be tumor suppressors, are downregulated while HOX family TFs are upregulated in GBM tissues, as compared to normal tissues. We also identified thousands of enhancers specifically active in GBM, as compared to normal cells, using H3K27ac ChIP-seq and ATAC-seq datasets. TF motif analysis of the identified enhancers revealed that motifs of Basic Leucine Zipper Domain (bZIP), CAAT box-binding transcription factor (CTF), homeobox (HOX), signal transducer and activator of transcription (STAT), and helix-turn-helix (HTH) family were enriched.
Conclusion: By integrating transcriptomic and epigenomic datasets, we identified and characterized TFs activated in GBM. We are currently investigating how ZFX cooperates with the TFs we identified as bound to the GBM-specific enhancers. These studies will provide insight into the molecular mechanisms underlying GBM.
Citation Format: Zexun Wu, Shannon Schreiner, Peggy J. Farnham, Suhn K. Rhie. Identification of transcription factors activated in glioblastoma multiforme [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2427.
Abstract 2428: NR4A1 ligands inhibits PSPC1 regulated EMT and stemness markers in breast, lung and liver cancer cells
Authors: Mohankumar Kumaravel , Shrestha Rupesh , Safe Stephen 
Published: July 2021

Abstract
The orphan nuclear receptor 4A1 (Nur77, TR3, NR4A1) is highly expressed in solid tumors and is a negative prognostic factor for survival of breast, lung, and liver cancer patients (1) and ongoing studies in our laboratory show that NR4A1 is pro-oncogenic in multiple tumor types. We have also identified a series of bis-indole derived compounds including 1,1-bis(3'-indolyl)-1-(p-hydroxyphenyl) methane (DIM-C-pPhOH; CDIM8) that act as NR4A1 antagonists that inhibit pro-oncogenic NR4A1-regulated genes and pathways in multiple cancer cell lines (2). Mechanistic studies show that many of the NR4A1-regulated genes involve interactions of NR4A1 with gene promoter DNA or with Sp-bound to GC-rich sequences and similar mechanisms have previously been observed for steroid hormone and other nuclear receptors. Our studies have identified paraspeckle component 1 (PSPC1) as an NR4A1 regulated gene in breast, lung and liver cancer cells and PSPC1 gene expression and protein levels are suppressed after treatment with C-DIM/NR4A1 antagonists. PSPC1 promotes tumorigenesis, epithelial-to-mesenchymal transition (EMT), stemness and metastasis and is a master activator of pro-metastatic switches and a potential target for anti-metastatic drugs. Knockdown of PSPC1 downregulates EMT and stemness markers in breast, lung and liver cancer cells. The novel buttressed analog of CDIM8 (DIM-C-pPhOH-3-Br-5-OCF3) at a concentration of 10 uM downregulates PSPC1 expression and its downstream targets (Snail, Slug, Nanog, OCT4, SOX2, c-myc). Knockdown of NR4A1 or SP1 inhibits PSPC1 expression in MDA-MB-231, H1299 and SNU449 cell lines respectively. These results identify PSPC1 as a novel gene that regulates EMT and invasion in these cancer cells and NR4A1 is an upstream regulator of PSPC1 gene expression. Thus, NR4A1 ligands act as an antimetastatic mimic by inhibiting PSPC1 and the mechanism of PSPC1 regulation involves ligand induced interactions of NR4A1/SP1 with a GC-rich region of the PSPC1 gene promoter.
References:1.Safe S, Jin UH, Hedrick E, Reeder A, Lee SO.
Minireview: Role of orphan nuclear receptors in cancer and potential as drug targets. Mol Endocrinol. 2014;28(2):157-72.
2.Mohankumar K, Li X, Sridharan S, Karki K, Safe S. Nuclear receptor 4A1 (NR4A1) antagonists induce ROS-dependent inhibition of mTOR signaling in endometrial cancer. Gynecol Oncol. 2019; 154(1):218-227.
Citation Format: Kumaravel Mohankumar, Rupesh Shrestha, Stephen Safe. NR4A1 ligands inhibits PSPC1 regulated EMT and stemness markers in breast, lung and liver cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2428.
Abstract 2430: HOXC6 promotes tumorigenesis of right-sided colon cancer
Authors: li Xin , Xia Jinhong , Ji Dengbo , Gu Jin 
Published: July 2021

Abstract
Colorectal cancer (CRC) is a common malignant tumor of the digestive tract, with high heterogeneity. The significant difference in prognosis and therapeutic response of left and right colon cancer is a hot spot in the field of colorectal cancer (CRC) in recent years. The prognosis and drug response of right-sided colon cancer were significantly worse than that of the left-sided colon cancer. Clinical trials have proved that MSI-H colon cancer is mainly seen in the right-sided colon cancer, and the prognosis is better than that of MSS. Because MSS and MSI-H patients have different molecular characteristics and prognosis, we searched for key genes of RCC and LCC in MSS and MSI-H groups respectively. The difference between the prognosis and the curative effect of the left and right-sided colon cancer is essentially the difference between the molecular pathology and the immune environment. Therefore, the aim of this study is to explore the molecular mechanism related to the prognosis of left and right-sided colon cancer in order to provide a molecular target for accurate treatment of left and right-sided colon cancer. First of all, bioinformatics methods were used to screen the right and left-sided CRC differentially expressed genes in MSS and MSI-H. The expression levels of these key genes in clinical samples and cell lines were detected by RT-qPCR and IHC, and their correlation with survival prognosis was analyzed. Homeobox C6(HOXC6)were selected. First, HOXC6 stable overexpressed and knockout colorectal cancer cell lines were established and its biological function was preliminarily studied, including transwell, wound-healing assay and CCK-8 tests. Finally, the ChIP-seq data were used to analyze the carcinogenic mechanism of HOXC6. Bioinformatics methods were used to screen 30 differentially expressed genes in MSS right and left CRC, of which HOXC6 in the right colon cancer was significantly higher than that in the left-sided colon cancer, which was positively correlated with TNM staging and lymph node metastasis. Kaplan-Meier survival analysis showed that the expression of HOXC6 was negatively correlated with the total survival of CRC patients (p=0.002). RT-qPCR and IHC showed that the expression of HOXC6 in the right-sided colon cancer was higher than that in the left, P&lt;0.05. Functional experiments showed that Hoxc6 promoted the growth, invasion and migration. ChIP-seq data and real-time PCR results showed that SPOCD1 and S100A1 were the target genes of HOXC6.
Citation Format: Xin li, Jinhong Xia, Dengbo Ji, Jin Gu. HOXC6 promotes tumorigenesis of right-sided colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2430.
Abstract 2435: Transcription Factor AP2 enhances malignancy of non-small cell lung cancer through upregulation of USP22 gene expression
Authors: Zhang Keqiang , Sun Ting , Li Wendong , Pangeni Rajendra , Raz Dan 
Published: July 2021

Abstract
Ubiquitin-specific peptidase 22 (USP22), a putative cancer stem cell marker, is frequently upregulated in various human cancers, and USP22 overexpression is associated with aggressive growth, metastasis, and therapy resistance in a number of human cancers including lung cancer. However, USP22 gene amplification seldom occurs, and the mechanism underlying USP22 upregulation in human cancers remains largely unknown. We set out to characterize and identify potential transcription factors that regulate USP22 expression in human non-small cell lung cancers (NSCLC). A luciferase reporter driven by the promoter region from -5085 to +55 of USP22 gene was constructed to screen a customized siRNA library targeting about 100 selected transcription factors that are expected to bind to the USP22 promoter. Luciferase reporter assays have revealed that knockdown of SP1 and activating transcription factor 3 (ATF3) promote USP22 expression, while knockdown of transcription factor 2α, 2β (TFAP2α and TFAP2β), cMyc, and PPAR α suppress USP22 expression. The specific binding of TFAP2α and TFAP2β to the USP22 promoter was further validated by binding site-directed mutagenesis and chromosome immunoprecipitation (Chip) assays. Furthermore, overexpression of TFAP2α and TFAP2β significantly up-regulate USP22 expression and downstream molecules of USP22, concurrently enhance the proliferation, migration and invasion of NSCLC A549 and H1299 cells in a partially USP22-dependent manner. In addition, TFAP2 was demonstrated to be associated with a poor prognosis of NSCLC patients. Moreover, TFAP2 protein expression correlated with USP22 protein expression in human NSCLC tissues. Taken together, the findings of this study indicate TFAP2 is an important transcription factor driving USP22 gene expression, and it promotes the progression of NSCLC partially through enhancing USP22 expression.
Citation Format: Keqiang Zhang, Ting Sun, Wendong Li, Rajendra Pangeni, Dan Raz. Transcription Factor AP2 enhances malignancy of non-small cell lung cancer through upregulation of USP22 gene expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2435.
Abstract 2437: Potency and selectivity evaluation of TEAD covalent inhibitors in living cells by two complementary mass spectrometry methods
Authors: Buzy Armelle , Paul Pascal , Hamon Stephanie , Venier Olivier , Guillemot Jean-Claude , Mikol Vincent , Moll Jurgen , Debussche Laurent , Valtingojer Iris 
Published: July 2021

Abstract
The HIPPO pathway plays an important role in cancer progression and among its essential components are the transcription factor proteins from the TEAD family. The TEAD family consists of 4 different members (TEAD1-TEAD4) who modulate gene expression through the interaction with co-activator proteins such as YAP1 and TAZ. Recent studies have shown that the central pocket of TEAD could be an alternative druggable approach for inhibiting YAP1. However, until recently, the irreversible binding mechanism of such a compound in living cells was not clearly established. Here, we report a mass spectrometry-based approach to demonstrate and quantitatively assess the target occupancy of TEADs covalent inhibitors both in vitro cellular and ex vivo tumor models. Applying our method to compounds from different chemical series confirmed that these inhibitors modulate the TEAD central pocket protein in cell cultures and led us to quantify their target occupancy for all TEADs expressed in our cellular models. We extended and strengthened the characterization of our inhibitors by implementing a proteomic LC-MS/MS cysteine profiling strategy - assessing the selectivity of inhibitors across free cysteine residues in the cellular proteome. As a result, we showed a high selectivity of the first inhibitors towards TEADs cysteine binding. To our knowledge, this is the first study proving by direct means and not by indirect biomarker and cell proliferation phenotypes that the central pocket of TEAD is actually available to ligand binding in cells, allowing direct measurement of target engagement for different TEAD proteins and finally demonstrating selectivity of inhibitors. These two-mass spectrometry-based methods developed here are very robust, powerful and can be currently used to help for the development of TEADs inhibitors
Citation Format: Armelle Buzy, Pascal Paul, Stephanie Hamon, Olivier Venier, Jean-Claude Guillemot, Vincent Mikol, Jurgen Moll, Laurent Debussche, Iris Valtingojer. Potency and selectivity evaluation of TEAD covalent inhibitors in living cells by two complementary mass spectrometry methods [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2437.
Abstract 2438: Genetic activation of JUN fuels the core regulatory circuitry of de-differentiated liposarcoma
Authors: Chen Ye , Dakle Pushkar , Tan Tuan Zea , Lin Ruby Yu-Tong , Puhaindran Mark E. , Gery Sigal , Xu Liang , Koeffler Phillip H. 
Published: July 2021

Abstract
Background: As a high-grade neoplasm of connective tissues, de-differentiated liposarcoma (DDLPS) is featured by genomic amplification of MDM2, CDK4 and JUN. Super-enhancer-driven master transcription factors (RUNX1, FOSL2 and MYC) establish a core feed-forward transcriptional circuit underpinning the DDLPS malignancy. However, an important question remains unanswered: how do genomic insults interact with epigenetic dysregulation in this disease?
Methods: We performed ChIP-seq analysis to map loading of transcription factors and histone marks on chromatin in DDLPS cells. Functional assays following genetic manipulation of target genes were conducted to explore their disease relevance.
Results: Genomic activation of JUN represents a promising prognostic marker for DDLPS. Meanwhile, JUN locus engaged excessive H3K27ac signals in both DDLPS tissues and cells, while a markedly lower level of H3K27ac deposition was observed in normal adipose tissues, adipocytes and MSCs. JUN formed a complex with FOSL2 and other co-factors in active chromatin, especially super-enhancers. Silencing of either JUN or its co-factors disrupted the core regulatory circuitry, which was accompanied by strong deleterious impacts on both cell viability and tumorigenicity. As BET bromodomain proteins dictate transcriptionally active chromatin, targeted degradation of BET proteins by PROTAC compounds exerted potent inhibitory effects on both JUN cistrome and DDLPS development.
Conclusion: Our study reveals that amplification and overexpression of JUN contribute to DDLPS development. DDLPS-associated genomic abnormalities collaborate with BET proteins to fuel disease-driving core regulatory circuitry.
Citation Format: Ye Chen, Pushkar Dakle, Tuan Zea Tan, Ruby Yu-Tong Lin, Mark E. Puhaindran, Sigal Gery, Liang Xu, Phillip H. Koeffler. Genetic activation of JUN fuels the core regulatory circuitry of de-differentiated liposarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2438.
Abstract 2447: TLR4 expression as a determinant of EMT and stress response gene expression in UV exposed human keratinocytes
Authors: Kirschnerova Viktoria , Vaishampayan Prajakta , Khawam Maria , Curiel-Lewandrowski Clara , Wondrak Georg T. , Dickinson Sally E. 
Published: July 2021

Abstract
Toll-like receptor 4 (TLR4) is gaining attention for its role in several cancer-associated signaling pathways, including the regulation of inflammation, DNA repair and epithelial-mesenchymal transition (EMT). There is evidence that TLR4 contributes to ultraviolet (UV) radiation-induced stress signaling in skin cells, including keratinocytes. Blockade of TLR4 activity using the pharmacological antagonist resatorvid (TAK-242) inhibits not only acute UV-induced signaling in vitro and in vivo, but blocks UV-induced skin carcinogenesis in mouse models. In order to genetically explore the role of TLR4 in keratinocyte responses to UV-induced stress, we recently utilized CRISPR/Cas9 techniques to delete TLR4 from the genome of human HaCaT keratinocyte cells in culture. First, using differential gene expression analysis (employing the EMTRT2 Profiler PCR array technology) it was observed that TLR4 KO status is associated with pronounced downregulation of EMT-related gene expression (MMP9, VIM, FN1, SNAI1, TWIST1) confirmed by independent RT-qPCR analysis. TLR4 was also a determinant of UV-induced inflammatory enzyme expression including PTGS2. Phenotypically, TLR4 KO cells displayed an attenuation of Matrigel transwell migration. TLR4 KO cells demonstrated increased sensitivity to UV-induced apoptosis as detected by flow cytometry of annexin V/propidium iodide stained cells, and by immunodetection of increased PARP cleavage. Moreover, NanoString nCounter expression analysis of murine skin chronically exposed to tumorigenic UV indicated that topical resatorvid treatment was associated with MMP9 downregulation. Future studies will employ this novel TLR4 KO keratinocyte cell line as a mechanistic tool to dissect the role of TLR4 in inflammatory signaling and EMT response in the context of skin photodamage and carcinogenesis. Supported by NIH grants ES029579, R03CA230830, P01CA229112.
Citation Format: Viktoria Kirschnerova, Prajakta Vaishampayan, Maria Khawam, Clara Curiel-Lewandrowski, Georg T. Wondrak, Sally E. Dickinson. TLR4 expression as a determinant of EMT and stress response gene expression in UV exposed human keratinocytes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2447.