bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1792: Exploring new therapeutic options for chemoresistant locally advanced lung cancer
Authors: Cuttano Roberto , Dama Elisa , Melocchi Valentina , Colangelo Tommaso , Carbonelli Cristiano , Maroni Giorgia , Levantini Elena , Bianchi Fabrizio
Published: August 2019

Background: Non-small-cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide. About one third of NSCLC patients present at diagnosis with locally advanced disease (stage IIIA) which are characterized by lymph node metastases. For this group of patients, neoadjuvant chemotherapy (NACT) followed by surgery is the best therapeutic option. However, clinical response to NACT is heterogeneous including patients with a complete eradication of lymph node metastases (pN0), and patients with persistent disease (pN2). Of note, 'pN0' patients showed a favourable prognosis (~50-60% survival, at 5-year) when compared to 'pN2' patients (~20-30% survival, at 5-year).

Methods: To identify new molecular determinants involved in NACT response, we recently performed whole-microRNA expression profiling of metastatic lung tumor cells, which were collected from mediastinal lymph nodes before NACT either by endobronchial ultrasound transbronchial aspiration (EBUS-TBNA) (n=11) or mediastinoscopy (n=41). In addition, we performed a meta-analysis of a cohort (n=84) of NSCLC (i.e. The Cancer Genome Atlas LUAD and LUSC dataset) with complete follow-up.

Results: We found a set of 20 miRNAs significantly differentially expressed in pN2 vs. pN0 tumors. Hierarchical clustering analysis using this 20-miRNA set revealed a clear separation of pN0 patients from pN2 patients. However, when the 20-miRNA set was used in the primary NSCLCs from TCGA dataset, the chemoresponsive and chemoresistant tumors could not be stratified. This possibly suggests unique properties of this 20-miRNA set and their modulated target genes in metastatic cells, and in the acquisition of a chemoresistant phenotype. Furthermore, by coupling miRNA and mRNA expression profile in metastatic samples, we rewired i) cancer intrinsic gene networks with a role in chemotherapy resistance, and ii) an augmented infiltration of tumor promoting immune cell populations (TILs).

Conclusions: Lung cancer metastases are intrinsically distinct and there exist molecular subtypes with different NACT response, identifiable by miRNA expression profile. Future efforts will be directed to identify the landscape of molecular mechanisms mediated by miRNAs, including the modulation of TILs possibly involved in the acquired chemoresistance.

Citation Format: Roberto Cuttano, Elisa Dama, Valentina Melocchi, Tommaso Colangelo, Cristiano Carbonelli, Giorgia Maroni, Elena Levantini, Fabrizio Bianchi. Exploring new therapeutic options for chemoresistant locally advanced lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1792.
Abstract 1794: A comparative microRNA expression analysis in breast cancer and melanoma tissues
Authors: Zhang Yanping , Peng Guangyong , Hsueh Eddy C.
Published: August 2019

Recently studies have revealed that a subset of microRNAs (miRNAs) is aberrantly expressed in the development and progression of a variety of cancers including breast cancer and melanoma. Several miRNAs have been implicated in tumorigenesis and progression of human cancer. However, the underlying mechanisms in different types of cancers remain largely unknown. The aim of the study was to compare the expression profiled of miRNA between breast cancer and melanoma patient’s tissues. Methods: Expression levels of 7 miRNAs (miR-15b, miR-142-3p, miR-142-5p, miR-142a, miR-199a, miR-221, miR-424, ) were determine in 17 cases of breast cancer and 25 melanoma tissue samples. MiRNA isolation from tissue samples were performed using Trizol reagent ( Sigma, USA). RNA quality and quantity were assessed using a BioMate TM 3 Series Spectrophotometers (Thermo, Madison, WI). Synthesis of cDNA with reverse transcriptase was performed by TaqManTM microRNA Reversed Transcription Kits, For analysis of miRNA expression, real-time q-PCR analyses were using TaqMan MicroRNA Assays. All real-time q-RT-PCR were performed on a 7300 real-Time PCR system (Applied Biosystems, USA). MiR-24 was used as the internal control for normalization. The relative miRNA expression levels were calculated using the 2 DΔCt methods. Values were presented as means ± standard deviation (SD). The comparison of miRNA levels in breast cancer and melanoma tissues were performed using Student’s t-test. A p value less than 0.05 was considered statistically significant. The expression of miRNAs in different breast cancer subtypes (infiltrating and Invasive ductal cancer) and melanoma (primary and metastatic) was also performed. Results: Relative miRNA expression levels (miR-15b, miR-142-3p, miR-142-5p, miR-146a, miR-199a, miR-221 and miR-424) were found to be differentially expressed in breast cancer and melanoma tissues. Significantly higher relative miRNA expression levels were detected in melanoma tissues compared with breast cancer tissues in all miRNAs evaluated except for miR-424. Four miRNAs (miR-15b, miR-146a, miR-221, and miR-143-3p) had relative expressions over 1.5 compared with only 1 miRNA (miR-15b) in breast cancer tissue. DΔCt values are plotted following comparison with endogenous levels of miR-24 assessed in each sample. Conclusions: Our study demonstrated that miRNA is not only a potential biomarker, but also a valuable therapeutic target for breast cancer and melanoma.

Citation Format: Yanping Zhang, Guangyong Peng, Eddy C. Hsueh. A comparative microRNA expression analysis in breast cancer and melanoma tissues [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1794.
Abstract 1799: MicroRNA hsa-miR-26a expression in hepatocellular carcinoma correlates with patient survival
Authors: Nakajima Go , Hayashi Kazuhiko
Published: August 2019

Background: MicroRNAs (miRNAs) are small noncoding RNAs that act as important regulators of gene expression at a post-transcriptional level. Using a next-generation sequencer system, in a previous study, we have identified 49 miRNAs expressed differentially by comparing 26 pair samples of hepatocellular carcinoma (HCC) tumor tissue and nontumor liver tissue. For this validation study, we selected hsa-miR-26a, one of the candidate miRNAs. We measured hsa-miR-26a expression in the validation set HCC samples using real-time quantitative polymerase chain reaction (qPCR) and examined the correlation between miRNA expression and clinical data. Patients and Methods: Overall, 40 patients with HCC who underwent tumor resection from 1991 to 1997 at the Department of Surgery, Institute of Gastroenterology, Tokyo Women’s Medical University, Japan were enrolled. Formalin-fixed paraffin-embedded samples were prepared using the standard protocol. The paraffin blocks were cut into 10-μm sections; HCC tumor and adjacent nontumor liver tissues were collected using laser-captured microdissection, and total RNA, including miRNA fractions, were isolated. Following the synthesis of a complementary DNA (cDNA) using an miRNA-specific primer, real-time qPCR was performed using an miRNA-specific primer/probe. Results: The hsa-miR-26a expression tended to be downregulated in HCC tumor tissue compared to nontumor liver tissue (P = 0.0799). Patients in the higher hsa-miR-26a expression group showed a significantly longer overall survival time compared with those in the lower hsa-miR-26a expression group (P = 0.0409). Conclusion: Inhibition of hsa-miR-26a expression might play a role in tumorigenesis, and hsa-miR-26a expression could be a potential prognostic factor for patients with HCC.

Citation Format: Go Nakajima, Kazuhiko Hayashi. MicroRNA hsa-miR-26a expression in hepatocellular carcinoma correlates with patient survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1799.
Abstract 1804: Role of microRNAs in colorectal adenoma progression
Authors: Wallace LaShanale M. , Chaudhry Aneese F. , Aikhionbare Karen I. , Yao Xuebiao , Aikhionbare Felix O.
Published: August 2019

Colorectal cancer (CRC) survival rates are stage related and majority of patients (75%) with CRC have sporadic of the disease which is in form of adenoma. Most of colorectal adenocarcinomas precursors are adenomatous polyps. Moreover, CRC appears to be an age related disease and studies have shown that mutations in mtDNA appear to accumulate with age because of inadequate repair. Also, oxidative stress has been shown to affect mitochondrial proteins resulting in a decrease of mitochondrial function associated with the precursor lesions of cancer. Recent findings suggest that the mitochondrion has sequences that code for certain miRNA which can regulate different pathways frequently dysregulated in cancer including colorectal cancer (CRC). Isolation of mitochondria RNA from colorectal precancerous polyps, cancer, and normal adjacent tissue samples was performed and assessed for gene expression differences using quantitative reverse transcription PCR. We demonstrate that mitochondria genome expression of miRNAs miR-24, miR 181, miR 210, and miR 21 in colorectal tissue samples varies during disease progression. Expression of microRNAs 24, 181, and 210 progressively increased from the earliest precancerous polyp to cancer. In addition, miR-210 and miR-181 expression increased 3 folds in villous adenomas when compared to tubular adenoma. Meanwhile, miR 21 increased progressively in adenoma tissues but decreased almost 4 folds in cancer tissues when compared to villous adenoma tissue. These results suggest miRNAs within the mitochondria genome may regulate important mitochondrial functional pathways leading to a more favorable environment for transformation or progression of adenomas to cancerous tumors.

Citation Format: LaShanale M. Wallace, Aneese F. Chaudhry, Karen I. Aikhionbare, Xuebiao Yao, Felix O. Aikhionbare. Role of microRNAs in colorectal adenoma progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1804.
Abstract 1809: LncRNA-SOX2-OT contributes to head and neck cancer cell proliferation by regulating DDIT4
Authors: Wang Ru , Yang Yifan , Tan Chen , Feng Ling , Fang Jugao
Published: August 2019

Background: Long non-coding RNAs (lncRNAs) have been shown to serve as an oncogenic role in human cancers. However, the role of lncRNAs in head and neck squamous cell carcinoma (HNSCC) is still unclear.

Methods and Results: Our previous microarray analysis showed that lncRNA SOX2-OT was upregulated in HNSCC tumor tissues and was correlated with DDIT4. To determine whether SOX2-OT served as a poor prognosis factor and targeted DDIT4 to facilitate cell proliferation and tumorigenesis in human HNSCC, qRT-PCR tested the relative expression of SOX2-OT and DDIT4 in 60 pairs of LSCC cancer tissues and the corresponding adjacent non-cancer tissues. Cell functional assay was conducted in vitro. We validated the up-regulation of SOX2-OT and DDIT4 in HNSCC tissues. Furthermore, we found that high SOX2-OT expression was significantly associated with advanced tumor differentiation, and worse prognosis. In vitro experiments demonstrated that SOX2-OT functioned as an oncogene by promoting cell proliferation and inhibiting cell apoptosis, and SOX2-OT targeted DDIT4 as its competing endogenous RNA in HNSCC cells.

Conclusions: Our findings for the first time identify the oncogenic role of SOX2-OT in HNSCC, demonstrating that SOX2-OT is a potential prognostic biomarker and promising therapeutic target in HNSCC.

Note: This abstract was not presented at the meeting.

Citation Format: Ru Wang, Yifan Yang, Chen Tan, Ling Feng, Jugao Fang. LncRNA-SOX2-OT contributes to head and neck cancer cell proliferation by regulating DDIT4 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1809.
Abstract 181: Prolyl-4-hydroxylase α subunit 3 promotes renal cancer progression and metastasis
Authors: Broome Larry E. , Wang Bingzhi , Zhao Menghuang , Yang Junhua , Bouamar Hakim , Cserhati Matyas , Sun Lu-Zhe
Published: August 2019

Collagen is the major component of the extracellular matrix, its overexpression has been shown to facilitate tumor development and progression. Prolyl-4-hydroxylase α subunit 3 (P4HA3) plays a crucial role in the synthesis of collagen, as it is involved in the catalysis to convert proline residues to 4 hydroxyproline residues, which are essential for the conformational stability of mature collagen. It has been reported that P4HA3 functions as a tumor promoter in gastric cancer. However, the role of P4HA3 in renal cell carcinoma (RCC) has not been established. To identify the role of P4HA3 in RCC, we analyzed the expression of P4HA3 in RCC patients and knocked down the expression of P4HA3 in two renal cancer cell lines with different P4HA3 siRNAs and P4HA3 shRNAs. We performed cell proliferation, migration and invasion assays in vitro and in vivo. Our results demonstrated that expression of P4HA3 was upregulated in RCC patients and knockdown of P4HA3 significantly inhibited renal cancer cell proliferation, migration and invasion. Mouse xenograft studies demonstrated the tumor promoting role of P4HA3 in renal cancer. Therefore, P4HA3 appears to be a potential therapeutic target for the treatment of RCC.

Citation Format: Larry E. Broome, Bingzhi Wang, Menghuang Zhao, Junhua Yang, Hakim Bouamar, Matyas Cserhati, Lu-Zhe Sun. Prolyl-4-hydroxylase α subunit 3 promotes renal cancer progression and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 181.
Abstract 1833: KLF4 regulates metabolic homeostasis in response to energy stress
Authors: Rosencrans William M. , Walsh Zachary H. , Blum Andrew C. , Hagos Engda G.
Published: August 2019

Krüppel-like factor 4 (KLF4) is a zinc finger-containing transcription factor with diverse regulatory functions in cellular growth, proliferation, differentiation, autophagy and genomic stability. Furthermore, KLF4 exerts a cell-cycle checkpoint effect in part by acting as a transcriptional activator of the cyclin-dependent kinase inhibitor, p21. Our lab has found that KLF4 regulates autophagy, mitophagy, and mitochondrial dynamics. Given the close relationship these processes have with cellular metabolism, we aimed to determine how KLF4 regulates glycolytic and oxidative metabolism. Using various assays, we find that cells lacking KLF4 have both oxidative and glycolytic impairments. Using western blotting, we find that when KLF4 is overexpressed in RKO human colorectal cancer cells, Hexokinase 2 (HK2), a rate determining enzyme in glycolysis, PKM2, a facilitator of lactate production, and MCT4, a lactic acid export protein and marker of glycolysis, are all significantly upregulated. In MEFs, we find that GLUT1, a glucose transport protein, has greater expression in WT than Klf4-null cells. Furthermore, we determined that KLF4 regulates aspects of the basal cell metabolism in a p21-dependent manner. Given that KLF4 has been found to act as an immediate early gene in response to certain stresses, and given the importance of fast metabolic shifts, we also look at how KLF4 influences short term metabolic stress responses. Since metabolic stress tests showed that WT MEFs have an increased glycolytic capacity, we investigate whether the cells have an increased ability to quickly intake glucose in response to particular stresses. Accordingly, we looked at GLUT1 expression and membrane localization with respect to KLF4. Immunostaining in RKO and MEFs suggests that KLF4 plays a role in localizing GLUT1 to the outer membrane, allowing for increased glucose intake. Taken together, our findings provide a rationale for the impaired metabolism of KLF4-null MEFs, where KLF4 plays a role responding to energy stress by modifying the localization of GLUT1, and regulating various metabolic enzymes both directly, and in a p21-dependent manner.

Citation Format: William M. Rosencrans, Zachary H. Walsh, Andrew C. Blum, Engda G. Hagos. KLF4 regulates metabolic homeostasis in response to energy stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1833.
Abstract 1835: Nqo1 ablation inhibits activation of the PI3K/Akt and MAPK/ERK pathways and blocks metabolic adaptation in hepatocellular carcinoma
Authors: Dimri Manali , Humphries Ashley , Laknaur Archana , Elattar Sawsan , Sharma Ashok , Kolhe Ravindra , Satyanarayana Ande
Published: August 2019

According to the World Health Organization, hepatocellular carcinoma (HCC) is the 2nd leading cause of cancer related deaths worldwide. HCC is highly resistant to conventional

chemotherapies; therefore, identification of specific molecular targets for the development of targeted therapies is indispensable. Cancer cells undergo metabolic adaptation to sustain uncontrolled proliferation and depends on the high rate of glycolysis and glutaminolysis. Hence, identifying and targeting proteins that assist cancer cells in metabolic reprogramming is an effective strategy to block their proliferation. We detected a strong upregulation of NAD(P)H Quinone Dehydrogenase 1 (Nqo1), a cytosolic flavoprotein, in vivo in mouse and human liver tumors as well as in liver cancer cell lines in vitro. Knockdown of Nqo1 in liver cancer cells by shRNA caused a significant downregulation of both glycolytic and glutaminolysis genes, impaired cell proliferation, colony formation and in vivo xenograft tumor growth. Nqo1-/- mice with HCC displayed significant reduction in tumor number, size and growth. In Nqo1+/+ mice, histological analysis of liver tumors revealed large tumor cells with hyperchromatic nuclei in a compact growth pattern, whereas Nqo1-/- livers appeared normal. Hyper-activated PI3K/Akt and MAPK/ERK signaling pathways play central role in cancer cell metabolic adaptation since their downstream effectors such as Akt and c-Myc control most of the glycolytic and glutaminolysis genes. Interestingly, at the molecular level, Nqo1 knockdown caused strong upregulation of the tumor suppressor PTEN, and enhanced activation of the serine/threonine phosphatase PP2A, leading to impaired activation of the PI3K/Akt and MAPK/ERK/c-Myc pathways. Our findings indicate that, by targeting PP2A and PTEN, Nqo1 enhances the expression and/or activity of PDK1, Akt, ERK1/2 and c-Myc and promotes cancer cell metabolic reprogramming and hence may function as a key therapeutic target for HCC inhibition.

Citation Format: Manali Dimri, Ashley Humphries, Archana Laknaur, Sawsan Elattar, Ashok Sharma, Ravindra Kolhe, Ande Satyanarayana. Nqo1 ablation inhibits activation of the PI3K/Akt and MAPK/ERK pathways and blocks metabolic adaptation in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1835.
Abstract 1853: Association of IDH genes with prostate cancer progression and their regulation by the androgen receptor
Authors: Gonthier Kevin , Poluri Raghavendra TK , Weidmann Cindy , Audet-Walsh Etienne
Published: August 2019

A primary function of the prostate is to synthesize and secrete high levels of citrate by way of a unique metabolic profile regulated by the androgen receptor (AR). Prostate cancer (PCa) is an androgen-dependent disease that is characterized by early reprogramming of citrate metabolism. Accordingly, mutations of the isocitrate dehydrogenase genes IDH1 and IDH2, which are key enzymes involved in the regulation of cellular citrate levels, have been demonstrated as important oncogenic events in several cancer types, including in about 3% of all PCa cases. However, if IDH1 and other IDH isoforms are associated with PCa progression as well as the regulatory factors controlling their expression remain mostly unknown. Across the human cancer spectrum, PCa appears to be the cancer type with the highest expression of IDH1, with levels even higher than common cancers associated with IDH1 mutations such as gliomas. Using publicly available datasets and quantitative PCR, we showed that IDH1 is the predominant IDH isoform expressed in PCa cells. In PCa, the androgen receptor was found to regulate several IDH isoforms in both in vitro and in vivo models of PCa, predominantly positively regulating IDH1. Chromatin immunoprecipitation experiments confirmed the recruitment of AR at several regulatory regions of IDH1 and IDH2. IDH1 and other IDH isoforms were shown to be significantly altered during PCa progression, which is consistent with a reprogramming of citrate metabolism in PCa. In addition, modulation of IDH expression significantly altered PCa cell proliferation and metabolism. Overall, our study indicates that IDH gene regulation is associated with PCa progression and that AR plays a significant role in the regulation of IDH genes.

Citation Format: Kevin Gonthier, Raghavendra TK Poluri, Cindy Weidmann, Etienne Audet-Walsh. Association of IDH genes with prostate cancer progression and their regulation by the androgen receptor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1853.
Abstract 1866: Sea cucumber (Stichopus japonicas) F2 enhanced TRAIL-induced apoptosis via XIAP ubiquitination-ER stress in colorectal cancer cells
Authors: Kim Jung lim , Park Seong Hye , Jeong Soyeon , Kim Bo Ram , Na Yoo Jin , Jo Min Jee , Jeong Yoon A , Yun Hye Kyeong , Kim Dae Yeong , Kim Bu Gyeom , Oh Sang Cheul , Lee Dae-Hee
Published: August 2019

Natural products have shown great promise in sensitizing cells to TNF-related apoptosis-inducing ligand (TRAIL) therapy. Sea cucumber (SC) extracts possess antitumor activity, and hence their potential to sensitize CRC cells to TRAIL therapy was evaluated. This study used Western blotting to evaluate the combination effects of SC and TRAIL in CRC and determined the molecular mechanism underlying these effects. SC fractions and TRAIL alone had no effect on apoptosis; however, combined treatment dramatically induced apoptosis of CRC cells, but not normal colon cells. Combined treatment induced expression of apoptotic proteins (poly (ADP-ribose) polymerase (PARP), caspase 3, and 8), and this effect was markedly inhibited by ubiquitination of X-linked inhibitor of apoptosis protein (XIAP). SC did not affect the mRNA levels, but increased proteasomal degradation and ubiquitination of XIAP protein. Furthermore, SC induced reactive oxygen species (ROS) production, thereby activating c-Jun N-terminal kinase (JNK) and endoplasmic reticulum (ER) stress-related apoptotic pathways in CRC. Taken together, our results demonstrate that the SC fraction may sensitize CRC cells to TRAIL-induced apoptosis through XIAP ubiquitination-ER stress.

Citation Format: Jung lim Kim, Seong Hye Park, Soyeon Jeong, Bo Ram Kim, Yoo Jin Na, Min Jee Jo, Yoon A Jeong, Hye Kyeong Yun, Dae Yeong Kim, Bu Gyeom Kim, Sang Cheul Oh, Dae-Hee Lee. Sea cucumber (Stichopus japonicas) F2 enhanced TRAIL-induced apoptosis via XIAP ubiquitination-ER stress in colorectal cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1866.
Abstract 1874: A ZEB1-regulated EMT transcriptional program is associated with early nodal metastasis and improved survival for stage N2/N3 breast cancer patients
Authors: Block C James , Wu Guojun
Published: August 2019

The epithelial-to-mesenchymal transition (EMT) is a transcriptionally-mediated process by which epithelial cells cease expression of cell-cell junction proteins, lose polarity, and begin expressing mesenchymal cytoskeletal and extracellular matrix components. While EMT is critical for normal development and wound healing, this process has been experimentally shown to be pathologically activated in cancer cells, leading to enhanced migratory and invasive capabilities, increased chemoresistance, and distant metastasis in mouse models. Consequently, EMT has been hypothesized to be a driving factor in metastatic progression and tumor recurrence in multiple cancer types, including breast cancer. EMT can be driven by several transcription factors (EMT-TFs) which are independently capable of inducing EMT and can regulate each other. Our group set out to identify EMT-TFs which were commonly activated across EMT models. By performing integrative analysis of multiple datasets, we determined that the EMT-TF ZEB1 demonstrates the most reproducible and consistent association with EMT in experimental models, breast cancer cell lines and in breast cancer patients. We then identified a high-confidence set of genes commonly upregulated in EMT and regulated by ZEB1. We observed that this ZEB1-associated transcriptional program is highly enriched for genes overexpressed in patients with early-stage node metastases compared to patients without lymph node involvement. Paradoxically, we observe that ZEB1 and ZEB1 associated genes exhibit higher expression in adjacent normal tissue samples than in primary tumor samples. Additionally, we discovered a reproducible association between ZEB1 and ZEB1-program gene expression and significantly improved recurrence-free and overall survival for breast cancer patients with distant lymph node involvement. This association was unique to ZEB1, as other EMT-TFs did not predict survival in these patients. In conclusion, our data demonstrate that the ZEB1-driven EMT program may contribute to early nodal metastasis, but that sustained ZEB1 expression may ultimately suppress cancer progression and improve survival. These findings may help clarify the specific contributions of different EMT-TFs to breast cancer progression.

Citation Format: C James Block, Guojun Wu. A ZEB1-regulated EMT transcriptional program is associated with early nodal metastasis and improved survival for stage N2/N3 breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1874.
Abstract 188: The oncometabolite d-2-hydroxyglutarate induces angiogenic activity through the vascular endothelial growth factor receptor 2 signaling pathway
Authors: Lee You Mie , Seok Jiyoon , Yoon Soo-Hyun , Lee Sun-Hee , Jung Jong Hwa
Published: August 2019

Mutation of isocitrate dehydrogenase (IDH) 1 (R132H) and IDH2 (R172K) and induction of hypoxia in various solid tumors results in changes to metabolic profiles, including production of the d- or l-forms of 2-hydroxyglutarate (2HG) from α-ketoglutarate in aerobic metabolism in the TCA cycle. However, it is unclear whether the oncometabolite d-2HG increases angiogenesis in endothelial cells. Therefore, in this study, we analyzed the levels of various metabolites, including d-2HG, under hypoxic conditions and in IDH2R172K mutant breast cancer cells by mass spectrometry. We then further evaluated the effects of this metabolite on angiogenesis in breast cancer cells. Our results showed that treatment with d-2HG increased the levels of secreted vascular endothelial growth factor (VEGF) in cancer cells and enhanced endothelial cell proliferation in a concentration-dependent manner. Wound healing and transwell migration were significantly increased by d-2HG to a level similar to that induced by VEGF. Tube formation was significantly stimulated by d-2HG, and chick chorioallantoic membrane angiogenesis was also enhanced by d-2HG. d-2HG activated VEGF receptor (VEGFR) 2 and VEGFR2 downstream signaling, extracellular signal-regulated kinase 1/2, focal adhesion kinase, AKT, and matrix metalloproteinase (MMP) 2. Taken together, our results suggested that d-2HG induced angiogenic activity via VEGFR2 signaling and increased MMP2 activity.

Citation Format: You Mie Lee, Jiyoon Seok, Soo-Hyun Yoon, Sun-Hee Lee, Jong Hwa Jung. The oncometabolite d-2-hydroxyglutarate induces angiogenic activity through the vascular endothelial growth factor receptor 2 signaling pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 188.
Abstract 1883: Ovarian cancer cells overcome the cytotoxic effect of cisplatin by hyper activating CCR9-mediated signaling in response to cisplatin
Authors: Mir Hina , Kapur Neeraj , Singh Shailesh
Published: August 2019

Ovarian cancer (OC) has highest mortality rate among the gynecological malignancies. Chemotherapies, in combination with surgery, are standard treatment offered to the patients with advanced OC. Although 80% patients initially respond to platinum-based chemotherapy, clinical responses remain short-lived due to chemoresistance. Most patients experience relapse and 50-60% of them die within 5 years. Therefore, development of novel therapies for OC remains a high priority. Hence, the major focus of this study is to define the biology of CCR9, which is highly expressed in OC in response to platinum drug. Effect of cisplatin on CCR9 expression and impact of CCR9 activation on molecular and cellular changes involved in overcoming the effect of cisplatin was determined by flow cytometric and western blot analyses. Our data show higher expression of CCR9 and CCL25 in OC cells in response to cisplatin treatment. Additionally, our data show higher expression and activation of estrogen receptor-α (ER- α) in OC cell after CCR9 activation. In addition to this, we show CCR9-mediated expression of Smoothened (SMO) known to support stem cell like phenotype by activating Hedgehog signaling and co-expression of CD44 and CCR9 as well as higher CD44 expression in OC cells following CCL25 treatment. CCR9-CCL25 axis promoted epithelial to mesenchymal transition. These observations strongly show an association of CCR9-CCL25 axis in OC pathogenesis, therapeutic resistance, and chemotherapeutic efficacy.

Citation Format: Hina Mir, Neeraj Kapur, Shailesh Singh. Ovarian cancer cells overcome the cytotoxic effect of cisplatin by hyper activating CCR9-mediated signaling in response to cisplatin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1883.
Abstract 1891: Oncogenic MDM2-containing exosomes promote pre-metastatic niche establishment
Authors: Casadei Lucia , Calore Federica , Deshmukh Ameya , Zewdu Abeba , Braggio Danielle A. , Fadda Paolo , Wabitsch Martin , Lopez Gonzalo , Leight Jennifer , Lev Dina , Croce Carlo M. , Pollock Raphael E.
Published: August 2019

Extracellular vesicles (EVs) have an important role in the communication between cells for their ability to transfer proteins and other biologically active molecules. EVs have been also described to have unique functions during pre-metastatic niche establishment and maintenance. MDM2 oncoprotein is the best-characterized cellular antagonist of p53 and its amplification is one of the most known mechanisms for tumor development. MDM2 amplification occurs in more than forty different types of malignancies and in 100% of well-differentiated and dedifferentiated liposarcoma (DDLPS). Retroperitoneal DDLPS is especially concerning given its propensity for multifocal local and distant recurrence, accounting for an overall survival rate of 10% at 10 years. Here, we show the presence of MDM2 in tumor EVs and specifically in both DDLPS cell line-derived EVs and patient-serum EVs. We also show the ability of MDM2-cargo to be transferred to recipient preadipocytes. Furthermore, we also demonstrated that this transfer is able to promote a pre-metastatic niche establishment in the DDLPS tumor microenvironment fat bearing areas, inducing: (I) tumor associated phenotype such as enhanced proliferation and migration, in the recipient preadipocytes, by downregulation of MDM2 downstream pathway; (II) pro-inflammatory phenotype through the release of pro-inflammatory cytokines such as IL-8; (III) release of active metalloproteinases such as MMP2 which is crucial for the invasiveness of many disseminating tumors. In conclusion here we report the presence of MDM2, in the patient circulation; importantly showing that MDM2 triggers the release of active MMP-2, we also propose a possible mechanism for pre-metastatic niche establishment and DDLPS multifocal loco-regional recurrence.

Citation Format: Lucia Casadei, Federica Calore, Ameya Deshmukh, Abeba Zewdu, Danielle A. Braggio, Paolo Fadda, Martin Wabitsch, Gonzalo Lopez, Jennifer Leight, Dina Lev, Carlo M. Croce, Raphael E. Pollock. Oncogenic MDM2-containing exosomes promote pre-metastatic niche establishment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1891.
Abstract 1917: High-throughput evaluation of treatment response in patient-derived glioma stem cell models
Authors: Zhang Ze-yan , Ding Yingwen , Ezhilarasan Ravesanker , Yang Jie , Long Lihong , Bronk Lawrence , Wang Qianghu , Sulman Erik P.
Published: August 2019

Although significant progresses in molecular oncology are being made using conventional cell lines, most therapies still fail in phase III clinical trials. Patient-derived models are being used more frequently as they are more faithfully representing the genomic features of primary tumors. However, one by one test of each model from large biobanks is extremely economy and time consuming. In this study, each of a panel of patient-derived glioblastoma stem cell (GSC) models was uniquely tagged by a lentiviral Cas9D10A and paired-gRNA targetable unique reporter (CAPTURE) barcoding system. Barcoded GSCs were then pooled evenly and following by radiation treatment (RT) in vitro. Amplicon sequencing was employed to count the barcodes distribution, which represent the relative cell number. The results showed that this approach faithfully identified the RT resistant GSCs from a mixing pool when comparing to the results from canonical clonogenic assay. In addition, a fluorescence marker will be switched by delivery of corresponding barcodes targeting CRISPR so that we can re-isolate interested cell models from the treated pool for investigating the treatment sensitivity and resistance mechanism. This study will provide a robust approach for therapeutic discovery take advantage of patient-derived models from large biobanks.

Citation Format: Ze-yan Zhang, Yingwen Ding, Ravesanker Ezhilarasan, Jie Yang, Lihong Long, Lawrence Bronk, Qianghu Wang, Erik P. Sulman. High-throughput evaluation of treatment response in patient-derived glioma stem cell models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1917.
Abstract 1969: Role of the CXCL17-CXCR8 (GPR35) axis in cutaneous squamous cell carcinoma
Authors: Khandelwal Alok R. , Alam Md Maksud , Moore-Medlin Tara , Savage Hillary A. , Nathan Cherie-Ann O.
Published: August 2019

Cutaneous squamous cell carcinoma (cSCC) is a keratinocyte-derived invasive and metastatic tumor of the skin and is the second-most commonly diagnosed form of skin cancer (16%) striking 200,000 Americans annually. Increased exposure to Ultraviolet-B (UVB) from solar radiation as a result of the depletion of the ozone layer and the widespread use of tanning beds are the primary reasons for the increased incidence of cSCC. Novel mechanism-based therapeutics are needed to prevent UVB-induced cSCC. Chemokines are a group of small chemotactic cytokines that can exert extensive and complex effects on cancer via interaction with their receptors. Chemokine (C-X-C motif) ligand 17 (CXCL17) is the latest member of the chemokine family, and recent studies have identified both a causative and suppressive role of CXCL17 and its cognate receptor GPR35 in tumorigenesis. The purpose of this study was to determine the role of CXCL17 and GPR35 in the pathogenesis of cSCC. CXCL17 and GPR35 were significantly overexpressed in cSCC cells (SCC12A and SCC118) compared to HaCaT cells. Stimulation with CXCL17 significantly induced cSCC cell proliferation, migration, and motility suggesting an essential role in tumor growth, invasion, and metastasis. Tumor-specific overexpression of CXCL17 was detected in human cSCC and UVB-induced mouse cSCCs that further bolsters our in vitro findings. Recent studies have also identified the role of CXCL17 in tumor cell infiltration and immune invasion leading to tumor progression. In our preliminary studies, increased expression of CXCL17 in cSCC correlated with an augmented number of CD3+, CD4+ and CD8+ T-cells infiltrates. Our data suggest that CXCL17 could potentially exert differential effects based on cell types in the tumor microenvironment. Our studies underscore the importance of CXCL17/GPR35 signaling in cSCC and provide a novel target for the treatment of non-melanoma skin cancer.

Citation Format: Alok R. Khandelwal, Md Maksud Alam, Tara Moore-Medlin, Hillary A. Savage, Cherie-Ann O. Nathan. Role of the CXCL17-CXCR8 (GPR35) axis in cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1969.
Abstract 1985: Reassessment of exosome composition
Authors: Jeppesen Dennis K. , Franklin Jeffrey L. , Higginbotham James N. , Zhang Qin , Coffey Robert J.
Published: August 2019

The heterogeneity of small extracellular vesicles and the presence of non-vesicular extracellular matter is a major obstacle to the study of exosomes. Here we employ two complementary methods, high-resolution density gradient fractionation and direct immunoaffinity capture, to demonstrate separation of small extracellular vesicles from non-vesicular material, and exosomes from other types of small extracellular vesicles. Cells secrete Argonaute 1-4 independently of exosomes. Extracellular RNA and RNA-binding proteins are differentially expressed between exosome and non-vesicle compartments. Exosomes do not possess a cytoskeleton and exclude glycolytic enzymes. Annexin A1 is a novel and specific membrane-associated protein marker of microvesicles shed directly from the cell plasma membrane, distinct from exosomes in vitro and in vivo. Small extracellular vesicles are not vehicles of active DNA release. Instead we propose a new model for active secretion of extracellular DNA through an autophagy- and multivesicular endosome-dependent but exosome-independent mechanism. A reassessment of exosome composition is necessary.

Citation Format: Dennis K. Jeppesen, Jeffrey L. Franklin, James N. Higginbotham, Qin Zhang, Robert J. Coffey. Reassessment of exosome composition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1985.
Abstract 1987: Exosome from CAF-ESCC aggravates tumor progression by its containing molecular miR-3656
Authors: Chen Xiaojia , Jin Yuan , Zhang Bihui , LI Xiaocen , Tian Baoqing , Wang Qiang , Hong An
Published: August 2019

Background: Exosome plays an important role in cell communication and cell environment. But the mechanism of how exosome derived from cancer-associated fibroblast (CAF) influent esophageal squamous cell carcinoma (ESCC) progression is not clear. In this work, compared with normal fibroblast-derived exosomes (NDEs), we found that CAF derived exosomes (CDEs) can enhance the growth, migration, and invasion of ESCC cells obviously. Therefore, we would like to clarify the role of CDEs in ESCC progression.

Methods and Results: Using microRNA array to analyze the different miRNAs in the CDEs and NDEs, we got a series of up-regulation miRNAs and two down-regulation miRNAs in CDEs. Among them, miR-3656 shows a significantly highest level. Then, we transfected miR-3656 mimics to the ESCC cells and found the mimics increased the growth, migration, and invasion of ESCC cells in cell proliferation assay. cell scratch tests and chamber invasion experiments. We established recombinant ESCC cells with miR-3656 high expression by the lentiviral system. The cells were injected into mice to form the xenograft tumor. The results showed the miR-3656 can enhance the tumor formation. After that, aided by TargetScan software, we found ACAP2, one of miR3656 targets, was significantly down-regulating in ESCC cells at the high level of miR-3656. When ACAP2 expression was knocked-down by siRNA, the proliferation and migration of ESCC cells was increased, which indicated ACAP2 a potential anti-tumor factor. Furthermore, we detected the different proteins between ESCC cells with high expression miR-3656 and normal ESCC cells through LC_MS; the results were analyzed by Integrate Pathway Analysis (IPA) system and many differential proteins were found to be involved of PI3K/AKT and Wnt pathways.

Conclusions: In this study, we found the CDEs promote the progression of their sounding ESCC cells in vitro and in vivo. The main reason is that CDEs carry some tumor-promoting miRNAs that influent the target ESCC cells. Especially, miR-3656 expresses the highest level of miRNAs in CDEs. It not only decreases the expression of ACAP2 but also impacts the expression of proteins involved PI3K/AKT and Wnt pathways in ESCC cells. The results indicate the CDEs playing a very important role in ESCC progression.

Citation Format: Xiaojia Chen, Yuan Jin, Bihui Zhang, Xiaocen LI, Baoqing Tian, Qiang Wang, An Hong. Exosome from CAF-ESCC aggravates tumor progression by its containing molecular miR-3656 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1987.
Abstract 1989: Breast cancer-derived exosomes modulate the endocytic pathway in brain endothelial cells
Authors: Morad Golnaz , Carman Christopher V. , Moses Marsha A.
Published: August 2019

Breast cancer brain metastasis is associated with high mortality and has shown a rising trend in incidence during the recent years. Given the rapid progression of brain metastasis, elucidation of the early events that lead to brain metastasis will pave the way to identifying potential diagnostic and therapeutic targets for early intervention. To explore the early mechanisms of breast cancer brain metastasis, we focused on the role of breast cancer-derived exosomes in this process. While extensive studies have explored the role of tumor-derived exosomes in tumor progression, the current understanding of the contribution of tumor-derived exosomes to brain metastasis remains limited. Using a mouse model of brain metastasis, we showed that pre-treatment of mice with exosomes derived from the brain-seeking MDA-MB-231 breast cancer cell line (Br-Ex) increases brain metastasis growth. This facilitation of brain metastasis was not observed with exosomes derived from the parental MDA-MB-231 cells (P-Ex). It is widely acknowledged that during early stages of brain metastasis, tumor cells grow along the brain vasculature, the blood brain barrier (BBB). To understand the mechanisms of the exosome-derived facilitation of brain metastasis, we studied the interactions between exosomes and the BBB. Using state-of-the-art models of the BBB and high-resolution microscopy, we have identified, for the first time, the mechanisms by which Br-Ex modulate the endocytic pathway in brain endothelial cells to decrease exosome degradation. Interestingly, our mechanistic studies showed that these effects are induced through the transfer of exosomal miRNAs enriched in Br-EX. Moreover, we have shown that Br-Ex can exclusively change the expression of integrins in brain endothelial cells in a way that it could alter the microenvironment around the BBB in favor of metastatic tumor cells. These findings indicate that exosomes derived from a brain-seeking subpopulation of breast cancer cells can exclusively modify the physiological regulation of the BBB at multiple levels to accelerate metastatic growth in the brain microenvironment. Taken together, these studies increase our understanding of the early events that facilitate brain metastasis and provide multiple potential targets for diagnostic and therapeutic applications in brain metastasis. (This work was supported by the Breast Cancer Research Foundation and NIH R01CA185530.)

Citation Format: Golnaz Morad, Christopher V. Carman, Marsha A. Moses. Breast cancer-derived exosomes modulate the endocytic pathway in brain endothelial cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1989.
Abstract 199: TGF-β in exosomes facilitates HNSCC progression by accelerating tumor angiogenesis
Authors: Ludwig Nils , Yerneni Saigopalakrishna S. , Hinck Cynthia S. , Pietrowska Monika , Hinck Andrew P. , Whiteside Theresa L.
Published: August 2019

TGF-β is a key regulator for tumor initiation and progression in head and neck squamous cell carcinoma (HNSCC). Tumor-derived exosomes (TEX) contain TGF-β and accumulate in the tumor microenvironment (TME). This study characterizes the TGF-β content of HNSCC-derived exosomes and evaluates in vitro and in vivo TGF-β signaling by exosomes that results in promotion of angiogenesis. TEX were isolated from supernantants of 5 different HNSCC cell lines by mini size exclusion chromatography (mini-SEC) and characterized by electron microscopy, nanoparticle tracking analysis and mass spectrometry (LC-MS/MS). TGF-β content in exosomes was evaluated by immunoblotting. Proliferation and migration of SVEC4-10 lymphendothelial cells in response to TEX were investigated in vitro and results were confirmed in vivo, using a matrigel plug model in mice. In these experiments a novel trivalent TGF-β receptor trap (mRER) was used to inhibit TGF-β signaling. TGF-β levels and activity were similarly measured in exosomes isolated from plasma of 20 HNSCC patients. TEX carried high levels of TGF-β and were found to be potent inducers of angiogenesis in vitro and in vivo through functional reprogramming and phenotypic modulation of endothelial cells. Proliferation (p<0.01) and migration (p<0.01) by SVEC4-10 were stimulated by TEX and effects were inhibited by mRER treatment of SVEC4-10 (p<0.05). TEX promoted formation of defined vascular structures in vivo, and increased (p<0.001) vascularization in matrigel plugs relative to control. Those effects were inhibited by mRER treatment (p<0.05). Exosomes in plasma of HNSCC patients carried varying levels of TGF-β, and patients with nodal metastases had elevated TGF-β levels (p<0.01) relative to patients with no meastasis.The data show that TGF-β signaling by TEX in HNSCC promotes angiogenesis and drives tumor progression. Future efforts should focus on silencing TEX, thereby adding new options to existing anti-angiogenic therapies.

Citation Format: Nils Ludwig, Saigopalakrishna S. Yerneni, Cynthia S. Hinck, Monika Pietrowska, Andrew P. Hinck, Theresa L. Whiteside. TGF-β in exosomes facilitates HNSCC progression by accelerating tumor angiogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 199.