bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4317: Application of gene methylation analysis for distinguishing primary tumor cells from normal epithelial cells from lung cancer patients in conditionally reprogrammed cell cultures
Authors: Wu Guodong , Xiao Yangui , Fang Fuyuan , Yao Da , Liu Ji , Cao Yanhua , Mao Yu , Yu Benjamin , Qian Youhui , Yu Derek
Published: August 2019

In vitro drug sensitivity assays utilizing “conditionally reprogrammed cells” (CRCs) from different primary tumor types to guide personalized therapy has been emerging. However, this approach has been hindered by the growth of normal epithelial cells in CRC cultures and the lack of effective biomarkers/assays to distinguish them from primary tumor cells. In this study, we developed a DNA methylation-based, real-time PCR assay to investigate the methylation status of SHOX2,PTGER4 and Septin9 gene promotor regions in human lung cancer formalin fixed paraffin-embedded (FFPE) samples. Our results showed increased DNA methylation for at least one of the three genes in 90% (28/31) of tumor samples as compared with the corresponding adjacent normal samples. Then we tested gene methylation in fresh surgical tumors versus normal adjacent tissues from lung cancer patients, and their respective in vitro CRC cultures. Increased methylation for at least one of the three genes was detected in 91.6% of tumor samples (11/12) as compared with the adjacent tissues, and these positive methylation statuses were maintained in 75% (9/12) of CRC cultures. Further results from in vitro assays showed a correlation between higher cell methylation status and increased growth property in 3D culture, as well as differential response to chemotherapy. Together, our results demonstrated that methylation of SHOX2, PTGER4 and Septin9 genes can be used as reliable biomarkers to monitor tumor cells in in vitro CRC cultures.

Citation Format: Guodong Wu, Yangui Xiao, Fuyuan Fang, Da Yao, Ji Liu, Yanhua Cao, Yu Mao, Benjamin Yu, Youhui Qian, Derek Yu. Application of gene methylation analysis for distinguishing primary tumor cells from normal epithelial cells from lung cancer patients in conditionally reprogrammed cell cultures [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 4317.
Abstract 4319: Hypoxia regulates m6A-reader YTHDF2 to fuel cancer-promoting inflammation in hepatocellular carcinoma
Authors: Hou Jiajie , Zhang He , Liu Jun , Zhao Zhenjun , Wang Jianye , Lu Zhike , Hu Bian , Zhou Jiankui , Zhao Zhicong , Feng Mingxuan , Zhang Haiyan , Shen Bin , Huang Xingxu , Sun Beicheng , Smyth Mark J. , He Chuan , Xia Qiang
Published: August 2019

Dynamic N6-methyladenosine (m6A) modification was previously identified as a ubiquitous post-transcriptional regulation that affected mRNA homeostasis. However, the m6A-related epitranscriptomic alterations and functions remain elusive in human diseases such as cancer. Here we show that hypoxia regulates the ‘reader’ protein YTH domain family 2 (YTHDF2), to ultimately stabilize the methylated oncogene mRNAs in inflammation-associated hepatocellular carcinoma (HCC). YTHDF2 silenced in human HCC cells or depleted in mouse hepatocytes evoked pro-inflammatory responses and accelerated tumor growth and metastatic progression. Under hypoxia, YTHDF2 processed the decay of m6A-containing interleukin 11 (IL11) and serpin family E member 2 (SERPINE2) mRNAs, which were responsible for the inflammation-mediated malignancy. Reciprocally, YTHDF2 transcription succumbed to hypoxia-inducible factor-2α (HIF-2α). Administration of a HIF-2α antagonist (PT2385) restored YTHDF2-programed epigenetic machinery and repressed HCC progression. Hence, our findings provide a new insight into the mechanism by which hypoxia adapts m6A-mRNA editing to promote cancer.

Significance: We show that hypoxia induces downregulation of m6A-reader YTHDF2 and hyper-upregulation of m6A-mRNAs in human HCC, which adapts m6A decoration into a cancer-promoting mechanism. Thus we identify YTHDF2 as a suppressor in tumor growth and metastasis by targeting IL-11 and Serpin E2.

Note: This abstract was not presented at the meeting.

Citation Format: Jiajie Hou, He Zhang, Jun Liu, Zhenjun Zhao, Jianye Wang, Zhike Lu, Bian Hu, Jiankui Zhou, Zhicong Zhao, Mingxuan Feng, Haiyan Zhang, Bin Shen, Xingxu Huang, Beicheng Sun, Mark J. Smyth, Chuan He, Qiang Xia. Hypoxia regulates m6A-reader YTHDF2 to fuel cancer-promoting inflammation 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 4319.
Abstract 4320: Enhanced expression of histone methyltransferases increase ERα expression in tamoxifen-resistant breast cancer cells
Authors: Kim Seungsu , Lee Min-Ho , Lee Mi-Ock
Published: August 2019

Estrogen receptor (ER)-positive breast cancers depend on ER signaling for growth and progression. Although tamoxifen remains the frontline treatment for such cancers, resistance to this drug limits its clinical efficacy. Nevertheless, the majority of tamoxifen-resistant patients retain ERα expression, providing a rationale for targeting ERα in these patients. Here, we found that the expression of mixed lineage leukemia (MLL) family proteins are increased in tamoxifen-resistant breast cancer cells and patients. Specifically, mixed lineage leukemia 3 (MLL3) and SET domain containing 1A (SET1A) depletion downregulated the expression of ERα protein and mRNA levels in a ER-positive breast cancer cell. Chromatin Immunoprecipitation analysis revealed that the knock-down of MLL3 and SET1A reduce the histone H3K4 methylation status in ESR1 promoter regions. We also observed the positive correlation between ERα level and MLL3/SET1A level in breast cancer clinical data sets. Importantly, MLL3 and SET1A depletion downregulated the ERα and its downstream growth-related protein levels, as well as the growth of both ER-positive and tamoxifen-resistant breast cancer cells. Likewise, MLL3 and SET1A knockdown decreased the proportion of cells entering the cell cycle. Additional fulvestrant treatment resulted in synergistic reduction of ERα levels and growth of the tamoxifen-resistant breast cancer cells. Thus, enhanced expression of MLL3 and SET1A in tamoxifen-resistant breast cancer cells supported ERα-dependent growth of breast cancer cells, suggesting that MLL3 SET1A might provide an attractive target for overcoming endocrine resistance.

Citation Format: Seungsu Kim, Min-Ho Lee, Mi-Ock Lee. Enhanced expression of histone methyltransferases increase ERα expression in tamoxifen-resistant breast 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 4320.
Abstract 4321: Epigenetic regulation of TP73 expression in HCC and GI cancer
Authors: Yao Zhixing , Poto Cristina Di , Ressom Habtom W. , Sherif Zaki A.
Published: August 2019

Tumorigenesis results from an accumulation of mutational and epigenetic changes that alter normal cell growth and survival pathways. TP73 is the homologue of the master tumor suppressor TP53 involved in cellular responses to stress and development. The TP73 gene encodes two different protein isoforms, TAp73 and ΔNp73, and maps to a region on chromosome 1p36 that is frequently deleted in neuroblastoma and other tumors, and is thought to contain multiple tumor suppressor genes. However, the analysis of p73-knockout mice yielded conflicting results with respect to tumor suppression. World-wide efforts in sequencing the TP73 gene in patient tumor samples have not provided evidence for genetic alterations as a common cause of TP73 inactivation in human cancer. The role of TP73 in tumorigenesis has remained elusive to date. In this study, we isolated two stem cell lines from normal young and old human liver tissues to determine TP73 expression in normal human liver stem cell and hepatocellular cancer (HCC) cell lines (HepG2, SNU398, SNU449 & SNU475), gastrointestinal cancer (GI) cell lines (Caco2 & HCT116) and a normal skin fibroblast cell line (HS27). The results show that TP73 only expresses in cancer cell lines. Further immunohistochemistry studies on human normal liver tissues as well as adjacent normal and cancerous liver tissues from HCC patients confirm that TP73 only expresses in the cancerous tissues. Moreover, methylation-specific PCR and bisulfite sequencing studies revealed that TP73 promoter is activated only in cancer cell lines by DNA methylation. Furthermore, ChIP assay results demonstrated that CTCF (a chromosomal networking protein CCCTC binding factor) binds to TP73 promoter and regulates TP73 expression. Our observation may prove significant for the development of future therapeutic and diagnostic applications.

Citation Format: Zhixing Yao, Cristina Di Poto, Habtom W. Ressom, Zaki A. Sherif. Epigenetic regulation of TP73 expression in HCC and GI 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 4321.
Abstract 4333: GCN5 positively correlates with c-MYC in non-small cell lung cancer
Authors: Mustachio Lisa Maria , Roszik Jason , Farria Aimee , Dent Sharon YR
Published: August 2019

Lung cancer causes the highest mortality in cancer-related deaths. Definition of novel molecular targets is needed since these cancers often become resistant to existing therapies. Epigenetic modifications may provide such targets. Recent reports suggest that the histone acetyltransferase (HAT) module within the transcriptional coactivator SAGA complex plays a role in cancer, creating a new link between epigenetic regulators and this disease. We aim to define the role of the SAGA HAT protein GCN5 in the regulation and function of oncogenes in lung cancers. Since prior work indicates that GCN5 regulates the stability of c-MYC and serves as a coactivator for MYC target genes, we hypothesize that there is a positive relationship between GCN5 and c-MYC in non-small cell lung cancer (NSCLC). We also propose that inhibition of GCN5 will reduce lung cancer formation and progression in mouse models of lung cancer driven by c-MYC overexpression. Our data indicate that both GCN5 and c-MYC proteins are upregulated in NSCLC cells compared to normal lung epithelial cell lines. This trend is observable only at the protein level, indicating that the upregulation of these proteins in NSCLC cells occurs post-transcriptionally. Both GCN5 and c-MYC expression levels are also positively associated in mouse and human NSCLC cell lines. Genetic repression of GCN5 in NSCLC cells reduces cell proliferation, increases the population of necrotic cells, and reduces c-MYC expression. Inhibition of the GCN5 active site using commercially available probes in NSCLC cells significantly (p < 0.05) reduces cell proliferation, increases the percentage of cells undergoing apoptosis, and reduces c-MYC expression. In human NSCLC tissues, GCN5 and c-MYC positively correlate and are associated with MYC target genes. Our ongoing studies will determine whether targeting GCN5 represses multiple oncogenic pathways in cancer and whether it inhibits oncoproteins difficult to directly target, such as MYC. We expect that our findings will highlight the importance of epigenetic regulators in carcinogenesis and extend beyond our current research focus to other cancer types.

Citation Format: Lisa Maria Mustachio, Jason Roszik, Aimee Farria, Sharon YR Dent. GCN5 positively correlates with c-MYC in non-small cell 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 4333.
Abstract 4334: Epigenetic reprogramming of tissue-specific transcription promotes metastasis
Authors: Teng Shuaishuai , Li Yang , Yang Ming , Qi Rui , Wang Qianyu , Lu Zhi , Wang Dong
Published: August 2019

Metastasis is the cause of death for 90% of cancer patients, but little is known about how cancer cells adapt to and colonize new tissue environments. Colorectal cancer (CRC) is the third most common cancer and the fourth most common cancer cause of death globally. Here, we investigated how metastatic cancer cells, such as CRC cells, with original-tissue specificity adapt to the environment of a distant tissue, like liver.

By analyzing transcriptome data of multiple cohorts of clinical samples and primary/metastatic cell lines, we found metastatic CRC cells lose their colon-specific gene transcription program and gain a liver-specific gene transcription program as they metastasize in the liver. The elevated expression of a number of liver-specific genes was confirmed in human liver metastatic tumors by immunohistochemistry. Through epigenomic profiling, we revealed this transcription reprogramming is driven by a reshaped epigenetic landscape of both typical and super-enhancers, and chemical inhibition of enhancer activity disrupts the ability of cells to execute this altered transcription program and consequently inhibits metastasis. Further we identified liver-specific transcription factors, FOXA2 and HNF1A, bind to the gained enhancers in liver metastatic cells and activate the expression of liver-specific genes. We also found both FOXA2 and HNF1A are highly expressed in human liver metastatic tumors compared to primary CRC tumors. Loss of function of FOXA2 or HNF1A inhibits liver-specific transcription and impairs the ability of metastatic CRC cells to adapt to liver. Consistently, the gain of function of HNF1A activated liver-specific transcription and enhanced CRC liver metastasis.

Our findings indicate the direct contribution of a reprogrammed tissue-specific transcription program, which is induced by reshaped epigenetic landscape as well as distant-tissue-specific transcription factors, to the adaption and colonization of metastatic CRC cells to a distal organ. Notably, in addition to CRC liver metastasis, this tissue-specific transcription reprogramming is also observed in other multiple distant organs and/or cancer types. In summary, our data suggest that epigenetically reprogrammed tissue-specific transcription promotes metastasis and might have implications for targeted anti-metastasis therapies.

Citation Format: Shuaishuai Teng, Yang Li, Ming Yang, Rui Qi, Qianyu Wang, Zhi Lu, Dong Wang. Epigenetic reprogramming of tissue-specific transcription promotes 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 4334.
Abstract 4339: Critical role of Dnmt3b catalytic activity in prevention of oncogene-induced leukemias and lymphomas
Authors: Lopusna Katarina , Nowialis Pawel , Haney Staci L. , Abraham Ajay , Opavska Jana , Opavsky Rene
Published: August 2019

Cytosine methylation is involved in a variety of biological processes, including development, X-chromosome inactivation, imprinting, and suppressing unwanted transcription. DNA methylation in mammalian cells is catalyzed by several DNA methyltransferases, including DNMT1, DNMT3A and DNMT3B. In addition to DNA methylation catalysis, these enzymes interact with a number of repressive proteins including HDACs, thereby repressing transcription in methylation-independent manner. The extent to which methylation-dependent and independent activities play roles in biological processes remains poorly understood. Here we examined the role of methyltransferase (MT) activity of Dnmt3b in normal mouse embryogenesis and in prevention of hematologic malignancies. Loss of Dnmt3b in mice is embryonically lethal between 10.5-12.5 dpc. Dnmt3b was also identified as tumor suppressor in murine models of lymphoid and myeloid malignancies. To address whether Dnmt3b’s catalytic activity is important for mouse embryogenesis and prevention of hematologic malignancies, we generated mice bearing a conventional Dnmt3bCI allele using CRISPR/Cas9 system. This catalytically inactive allele of Dnmt3b (Dnmt3bCI) was generated by a double amino acid substitution in enzyme active center (P656V and C657D) abolishing its ability to perform transfer of methyl group onto cytosine. Surprisingly, Dnmt3bCI/CI mice were born at similar ratios as wild-type littermates and lived over 12 months suggesting that Dnmt3b is dispensable for both pre- and post-natal development. Importantly, MYC-induced T-cell lymphomagenesis was accelerated in the absence of Dnmt3b’s methyltransferase activity in MYC;Dnmt3bCI/CI mice. Global gene expression and methylation profiling revealed a number of deregulated events specific for MYC;Dnmt3bCI/CI, resulting in activation of Pak, Fgfr and Igf2 signaling and downregulation of tumor suppressors (e.g. Lrp12, Dusp6 and Marcks). The lack of Dnmt3b catalytic activity also accelerated a development of acute myeloid leukemia induced by MLL-AF9 overexpression in mice. Altogether, our data suggest that Dnmt3b’s MT activity is dispensable for mouse development but critical to prevent hematologic malignancies.

Citation Format: Katarina Lopusna, Pawel Nowialis, Staci L. Haney, Ajay Abraham, Jana Opavska, Rene Opavsky. Critical role of Dnmt3b catalytic activity in prevention of oncogene-induced leukemias and lymphomas [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 4339.
Abstract 4349: Predicting gene expression from plasma cell-free DNA using both the fragment length and fragment position
Authors: John John A. St , Gafni Erik , White Brandon , Kannan Ajay , Hansen Loren , Jaroszewicz Artur , Kundaje Anshul , Boley Nathan
Published: August 2019

The ability to use a blood sample to determine the transcriptional state of cells that are releasing DNA into the bloodstream of a patient may be helpful in a variety of clinical applications. Here we present a case study of a gene expression prediction model that uses cell-free DNA (cfDNA) fragment coverage data generated by high-throughput sequencing to predict which genes are highly or lowly expressed in the cells contributing to that cfDNA. We evaluated a number of models, including a convolutional neural network that takes cfDNA fragment information (the density of both fragment midpoint and length by genomic position) over a transcription start site (TSS) as input, and outputs a predicted probability of whether that gene is highly expressed in cfDNA-producing cells. When we trained the convolutional model on a set of 554 genes with TSSs that were either constitutively expressed or unexpressed across leukocyte samples from the NIH Roadmap Epigenome Mapping Consortium, we achieved ~0.97 AUC in cross validation. With other models and splits of the data, we observed AUCs ranging from 0.95 to 0.99 on this gene-expression task. Next, we were interested in whether this trained model could answer specific clinical questions. For example, we hypothesized that we should see an increased influence of colon gene expression profiles in colorectal cancer patients with a higher fraction of circulating tumor DNA. To test this hypothesis, we applied our models to a set of genes with colon-specific expression, which generated a list of probabilities of each gene being expressed in each sample. We then applied simple models on the these lists of probabilities to predict whether a patient had CRC or was healthy. This yielded cross validation AUCs between 0.85 and 0.95 across many of the models we tested in differentiating healthy patients from colorectal cancer patients with tumor fraction over 5%. These results suggest a path forward for modeling transcriptional states using cfDNA sequencing data, which will enable greater insights from cfDNA that could augment those provided by other analytes.

Citation Format: John A. St John, Erik Gafni, Brandon White, Ajay Kannan, Loren Hansen, Artur Jaroszewicz, Anshul Kundaje, Nathan Boley. Predicting gene expression from plasma cell-free DNA using both the fragment length and fragment position [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 4349.
Abstract 4350: MAPK1
E322K somatic mutation promotes cell proliferation via positive feedback regulation of EGFR/RAF signaling
Authors: Zhang Jianqing , Brooke Dewey J. , Blair Jessica P. , Riby Jaques , Ojesina Akinyemi I.
Published: August 2019

The RAS-MAPK signaling cascade serves as the central node in the transduction of signals starting from the binding of the epidermal growth factor (EGF) ligand to the membrane receptors (EGFR) through to the nucleus. Recent studies suggested that different defined thresholds of RAS-MAPK activity are required for maintaining homeostasis as opposed to facilitating malignant transformation, and that Ras-oncogenic toxicity is quantitatively dictated by MAPK function. We and others have demonstrated that somatic MAPK1(E322K) mutations in MAPK1 occur in 8% of cervical squamous cell carcinoma (CSCC) and 3% of head and neck squamous cell carcinoma (HNSCC), suggesting that mutant MAPK1 may play an important part in tumorigenesis. To investigate the molecular mechanism involved in the effects of the mutated MAPK1, we transduced MAPK1 wild type and MAPK1(E322K) mutant as lentiviral constructs into the untransformed human embryonic kidney cell line HA1E. We found that cells expressing MAPK1(E322K) exhibited higher rates of proliferation and cellular mobility compared to the cells transduced with wild type MAPK1. However, the cells expressing wild type MAPK1 presented poor cell proliferation compared to the cells transduced with the empty lentiviral vector. Western blot analyses showed approximately two-fold increase phosphorylation of ERK (T202/Y204) following EGF treatment in the cells harboring the mutation compared to the cells expressing the wild type MAPK1. A similar pattern was observed with phospho-EGFR (T669, Y1148, Y1068, Y1148) phospho-cRaf (S259, S338, S289/296/301), phosphor-RSK1, phospho-AKT (S473, T389), phosphor-GSK3β (S9/21) and phospho-STAT5 (Y694), suggesting that MAPK1(E322K) mutation might reinforce the positive/negative feedback loop on EGFR and cRaf. Our data contributes to the understanding of the roles of MAPK1 in the RAS/RAF-MEK-MAPK1 signaling pathway and might help identifying possible therapeutic targets for HNSCC and CSCC patients with tumors harboring somatic MAPK1(E322K) mutations.

Citation Format: Jianqing Zhang, Dewey J. Brooke, Jessica P. Blair, Jaques Riby, Akinyemi I. Ojesina. MAPK1E322K somatic mutation promotes cell proliferation via positive feedback regulation of EGFR/RAF signaling [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 4350.
Abstract 4370: Knockdown of PKM2 induces autophagic cell death via Akt/mTOR pathway in human prostate cancer cells
Authors: Dey Prasanta , Kundu Amit , Lee Byung Mu , Kim Hyung Sik
Published: August 2019

Pyruvate kinase M2 (PKM2) is crucial for aerobic glycolysis that is highly expressed in the various cancer tissues. Although high expression of PKM2 is observed in prostate tumor tissues, its functional role against affecting cancer metabolism is not clearly understood. Herein, we investigate its role in regulating autophagy and its associated pathways. In the current study, various PKM2-siRNA constructs were used to knockdown the PKM2 expression and observed its cellular pathways on autophagy. Cell viability was significantly reduced in PKM2-siRNA transfected prostate cancer cells, DU145. Acridine orange staining and immunoblotting analysis showed that downregulation of PKM2 markedly increased autophagy cell death. In addition, immunoblotting analysis exhibited the blocking the protein kinase B (Akt) and mTOR1 pathways, which subsequently down-regulated the expression of glycolytic enzymes lactate dehydrogenase A (LDHA) and Glucose transporter 1 (GLUT1). To the best of our knowledge, this is the first study that inhibition of PKM2 changes in cancer cell metabolism and induces autophagy, thus, providing new perspectives into the mechanism of its anticancer activity against prostate cancer, DU145.

Citation Format: Prasanta Dey, Amit Kundu, Byung Mu Lee, Hyung Sik Kim. Knockdown of PKM2 induces autophagic cell death via Akt/mTOR pathway in human prostate 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 4370.
Abstract 4373: Lack of FGF21 promotes NASH-HCC transition via exosome-mediated carcinogenetic signaling
Authors: Yu Youxi , Martin Robert C. , Zheng Qianqian , Shi Xiaoju , Liu Xingkai , Guo Wei , Li Suping , Zhang Ping , Li Yan
Published: August 2019

Background: Nonalcoholic steatohepatitis (NASH) is the most severe form of non-alcoholic fatty liver disease (NAFLD) and a potential precursor of hepatocellular carcinoma (HCC). FGF21, with its metabolic benefits, was reported as the liver safeguard to protect from obesity, diabetes, and NAFLD. Recent study showed an exosome based FGF21 inhibition via organs communication to regulate hepatic metabolism. However, the role of the exosome-FGF21 axis in NASH-HCC has never been studied.

Aim: To study the exosome based FGF21 metabolic regulation during NASH-HCC carcinogenetic transformation.

Methods: NASH-HCC model was induced in FGF21 knockout (KO) mice and wild type (WT) control using methionine-choline-deficient diets (MCD) and diethylnitrosamine (DEN). The exosomes releasing, miRNA and protein contents were analyzed in the primary cultured hepatocytes from the liver tissues of NASH-HCC mice (KO and WT) as well as benign controls. FGF21 was knockdown (KD) by shRNA in a mouse hepatoma cell line (Hepa1-6) and a benign mouse hepatocyte cell line (FL83B) to investigate the potential carcinogenetic signaling related to exosome-FGF21 axis.

Result: NASH-HCC model was successfully established with confirmation of the metabolic disorders and liver cancer development. Early HCC detection and increased HCC incidence were found in the KO-MCD mice (P<0.05 versus KO-CD and WT controls). The collected exosomes from primary cultured hepatocytes and cell lines were identified using the antibodies of Tsg101, CD63 and cytochrome C by western blotting and immunofluorescent staining. In the KO-MCD mice, exosomes retained in the cytosol of benign hepatocyte but released increasingly from cancer cell. Similar results were showed in the Hepa1-6KD cells and the FL83BKD cells. Aberrant EMT, and p53 and Wnt signaling was found in the KO-MCD mice as well as Hepa1-6KD cells challenged with free fatty acid.

Conclusion: lack of FGF21 plays critical role for the exosome retention in hepatocytes of NASH mice but not in HCC mice. Abnormal traffic of exosomes contribute to the carcinogenetic signaling. This work was supported by an Institutional Development Award (IDeA) from the NIGMS of the National Institutes of Health under grant number P20GM113226.

Citation Format: Youxi Yu, Robert C. Martin, Qianqian Zheng, Xiaoju Shi, Xingkai Liu, Wei Guo, Suping Li, Ping Zhang, Yan Li. Lack of FGF21 promotes NASH-HCC transition via exosome-mediated carcinogenetic signaling [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 4373.
Abstract 4390: Dietary soy isoflavone equol promotes breast cancer progression via regulation of protein synthesis initiation
Authors: Cruz-Collazo Ailed M. , Parra Columba De la , Schneider Robert , Dharmawardhane Suranganie
Published: August 2019

We previously reported that dietary daidzein, a soy isoflavone, increased breast cancer progression in a nude mouse model of breast cancer metastasis. Further studies indicated that equol, a metabolite of the soy isoflavone daidzein is responsible for the cancer promoting effects of soy isoflavones in breast cancer cells in vitro. We reported that equol increases the expression of the eukaryotic protein synthesis initiation factor eIF4G1, and the transcription factor c-Myc. Increased eIF4G in response to equol resulted in the non-canonical protein synthesis of pro-cancer molecules such as Cyclin D, Bcl-XL, p120, matrix metalloproteinases, etc., as confirmed by ribosome profiling and eIF4G1 knockdown studies. This study tested the hypothesis that equol promoted breast cancer progression via eIF4G1 upregulation. Immunocompromised nude mice were used to establish mammary fatpad tumors from human metastatic breast cancer cells expressing eIF4G1 knockdowns via a Tetracycline-inducible promoter. Oral gavage of 50 mg/kg BW Doxycycline 3X a week for 4 weeks was enough to knockdown eIF4G1 expression by ~80%. Therefore, one week following tumor establishment, the mice were treated with vehicle (90% corn oil, 10% ethanol), equol, Doxycycline or, equol and Doxycycline. Data show that as expected, equol treatment increased tumor growth relative to vehicle treatment. Moreover, eIF4G1 knockdown abolished the effect of equol on tumor growth. Interestingly, the effect of eIF4G1 knockdown on mammary tumor growth was not evident until 55 days of doxycycline treatment. Therefore, we tested the cell and tumor lysates for potential expression of the eIF4G1 isoform Dap5 when eIF4G1 is knocked out. Dap5 protein was expressed in eIF4G1 knockdown cells and was increased by equol treatment, indicating that Dap5 may compensate for eIF4G1 knockdown by inducing alternative cap-mediated protein synthesis. In conclusion, the cancer promoting effect of equol is abolished in breast tumors where eIF4G1 expression was stably knocked down, thus, validating our hypothesis that equol increases breast cancer progression via upregulation of eIF4G1 dependent protein synthesis initiation.

Citation Format: Ailed M. Cruz-Collazo, Columba De la Parra, Robert Schneider, Suranganie Dharmawardhane. Dietary soy isoflavone equol promotes breast cancer progression via regulation of protein synthesis initiation [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 4390.
Abstract 4401: Implications of STAG2 loss in Ewing sarcoma
Authors: Gorthi Aparna
Published: August 2019

Ewing sarcoma is an aggressive pediatric bone and soft tissue cancer that is driven by a translocation event, predominantly involving the RNA binding protein EWSR1 and the transcription factor FLI1. The resultant fusion transcription factor EWSR1-FLI1 is a potent oncogene and drives dramatic changes in the transcriptional landscape of Ewing sarcoma. Our previous work demonstrated that EWS-FLI1 drives aberrant accumulation of R-loops, increases replication stress and suppresses homologous recombinational repair in Ewing sarcoma. Similar phenotypes were also observed with the loss of wildtype EWSR1. However, surprisingly, Ewing tumors have highly stable genomes. The most common mutation is in the STAG2 gene, a component of the cohesin ring complex.

It was recently reported that STAG2-cohesin complexes not only participate in maintaining chromatin architecture but also play a key role in directing tissue-specific gene expression by interacting with transcription factors. In the present study, we evaluate the STAG2 function in R-loop metabolism, replication and repair and attempt to identify the reason for its selective loss in Ewing sarcoma. Using biochemical and biophysical experimental strategies, we observed that STAG2 and EWSR1 both bind R-loop structures with high affinity and are protective against RNaseH-mediated degradation of R-loops to significantly different degrees. We also examine the effect of STAG2 loss on R-loop accumulation and replication stress in Ewing and non-Ewing cell lines. Finally, we evaluate the correlation between STAG2 binding sites and R-loops. We propose that loss of STAG2 alleviates some of the genomic stresses experienced in Ewing sarcoma.

Note: This abstract was not presented at the meeting.

Citation Format: Aparna Gorthi. Implications of STAG2 loss in Ewing sarcoma [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 4401.
Abstract 4404: E-p-Methoxycinnamoyl-α-l-rhamnopyranosyl ester (MCR) increases Nrf2 stability by inhibiting ubiquitination in HaCaT cells
Authors: Jeong Jiwon , Wahyudy Lilik D. , Yang Heejung , Kim Jung-Hwan
Published: August 2019

The nuclear factor erythroid-derived 2-related factor 2 (Nrf2) is a key regulator of gene expression during oxidative stress and drug detoxification. Thus, identifying Nrf2 activators to protect from possible cell damage is necessary. In this study, we investigated whether E-p-methoxycinnamoyl-α-l-rhamnopyranosyl ester (MCR), a phenylpropanoid isolated from Scrophularia buergeriana, can activate Nrf2 signaling in human keratinocytes (HaCaT). First, we determined the dose- and time-dependent effects of MCR on the expression and activity of Nrf2. The antioxidant response element-luciferase reporter assay and western blot analysis results showed that MCR markedly induced Nrf2 activity and its protein expression, respectively. Further, MCR increased both the mRNA and protein levels of heme-oxygenase-1, one of the Nrf2 target genes, in the cells. Interestingly, we found that Nrf2 stability was remarkably enhanced by MCR. Furthermore, ubiquitin-dependent proteasomal degradation of Nrf2 was significantly reduced by MCR. Thus, MCR might afford skin protection by enhancing Nrf2 stability or by blocking its proteasomal degradation.

Citation Format: Jiwon Jeong, Lilik D. Wahyudy, Heejung Yang, Jung-Hwan Kim. E-p-Methoxycinnamoyl-α-l-rhamnopyranosyl ester (MCR) increases Nrf2 stability by inhibiting ubiquitination in HaCaT 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 4404.
Abstract 4405: Acetylation of KLF5 controls the cell fate of basal progenitors during postnatal development and regeneration of mouse prostates
Authors: Zhang Baotong , Dong Jin-Tang
Published: August 2019

Adult stem cells are responsible for lineage specification in epithelial homeostasis, and thus are essential for both postnatal development and tissue regeneration. Genetic changes in adult stem cells have been revealed to promote cancer development. Dissecting the underlying regulators is critical for understanding the intrinsic mechanisms that regulate adult stem cell function, yet whether PTM of a single molecule determines cell fate of adult stem cells during organ development has not been well established, particularly for glandular organs. Using the androgen-dependent mouse prostate as an experimental system, an ideal system for understanding epithelial hierarchy, we examined the Krüppel-like factor 5 (Klf5) transcription factor and its acetylation for their roles in postnatal development and regeneration. The absence of KLF5 eliminated basal progenitor activities in both human and mouse prostate epithelial cells, as indicated by a compromised sphere and organoid formatting capabilities, downregulation of basal cell markers, and decreased basal cells. Klf5 was also necessary for basal progenitor cells to maintain their luminal differentiation capacity, as loss of Klf5 also decreased basal progenitor-derived luminal cells and their proliferation in mouse prostates. Interestingly, the progenitor activity of Klf5 was restricted to deacetylated Klf5 (deAc-Klf5), as indicated by in vitro and in vivo assays. On the other hand, acetylated Klf5 (Ac-Klf5) was essential for proper formation and maintenance of luminal structures in mouse prostates, as deacetylation of Klf5 interrupted the formation of normal luminal structures, almost eliminated the castration resistance in basal progenitor-derived luminal cells, and attenuated the luminal regeneration after a castration-regeneration cycle, even though deAc-Klf5 promoted basal-to-luminal differentiation and the proliferation of luminal cells during postnatal development. Mechanistically, deacetylation of Klf5 increased Notch signaling activity. These findings indicate that Klf5 and its acetylation control the cell fate of prostatic basal progenitors, which have a distinct contribution to postnatal development and regeneration of the prostate. Considering that androgen deprivation therapy (ADT) is currently a major therapeutic strategy in the treatment of prostate cancer, our findings also provide a potential mechanism for prostate cancer progression, i.e., interruption of Klf5 acetylation could sensitize castration-resistant prostate cancer to ADT or slow the recurrence of prostate cancer after ADT.

Citation Format: Baotong Zhang, Jin-Tang Dong. Acetylation of KLF5 controls the cell fate of basal progenitors during postnatal development and regeneration of mouse prostates [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 4405.
Abstract 4410: Predictive model of palbociclib response reveals indications in which CDK4/6 inhibitor can be a potential combination partner
Authors: Hafner Marc , Dompe Nicholas , Zhou Wei , Lin Eva , Durrbaum Milena , Daemen Anneleen , Junttila Melissa R. , Metcalfe Ciara , Merrick Karl
Published: August 2019

CDK4/6 inhibitors are a promising class of targeted therapies, with three molecules (palbociclib, ribociclib, and abemaciclib) approved for advanced hormone-receptor positive (HR+) breast cancers. With hope of building on this success, clinical trials across multiple oncology indications have been initiated. The determinants of response to CDK4/6 inhibitors, beyond pRb-deficiency and Cyclin-E amplification, remain unclear as limited clinical data have been published in indications other than HR+ breast cancers. Here, we measured the response to palbociclib across a panel of tumor cell lines from multiple indications and were able to build a computational model predictive of drug response using the expression of a handful of transcripts. This model revealed cellular states that are related to palbociclib resistance, such as G1-S entry deregulation, or sensitivity, such as competent p53 pathway and high KRAS signaling.

As validation of our model, we compared its predictions with dependency scores from Project Achilles and found good correlations between predicted sensitivity and dependence on CDK4 and CCND1. By applying the predictive model on TCGA data, our analysis uncovered indications, disease subtypes, and genomic alterations that are related to the predicted response to palbociclib in addition to Rb-deficiency and Cyclin-E amplification. As expected, HR+ breast cancer was predicted to be among the most sensitive diseases, along with prostate, thyroid, and some renal cancers. Among non-small cell lung cancers, squamous-cell tumors are predicted to be more resistant than adenocarcinomas. In adenocarcinomas, BRAF and KRAS mutations are predicted to be associated with increased responsiveness. Combining these results with published biomarkers of resistance to standard of care therapies reveals indications in which CDK4/6 inhibitors can be potential combination partners. These results may guide the design of new trials for CDK4/6 therapies and the evaluation of better stratification biomarkers.

Citation Format: Marc Hafner, Nicholas Dompe, Wei Zhou, Eva Lin, Milena Durrbaum, Anneleen Daemen, Melissa R. Junttila, Ciara Metcalfe, Karl Merrick. Predictive model of palbociclib response reveals indications in which CDK4/6 inhibitor can be a potential combination partner [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 4410.
Abstract 4411: Identification of novel natural compounds as potential inhibitors of cyclin dependent kinases
Authors: Sankaranarayanan Ranjini , Dachineni Rakesh , Kesharwani Siddharth , Dhandapani Ramesh Kumar , Tummala Hemachand , Gunaje Jayarama B.
Published: August 2019

Colorectal cancer (CRC) is the second highest cause of cancer deaths in the United States according to CDC (2017). Lifestyle factors including, but not limited to, elevated Body Mass Index (BMI), obesity and diet are proposed to lead to the development of CRC with an estimated annual incidence of 95,520 cases. Increasing evidences suggest that a diet rich in fruits and vegetables will reduce incidences of colorectal adenomas. We hypothesize that the natural compounds present in fruits and vegetables, along with their metabolites generated by the gut microbiota, could be responsible for cancer prevention by targeting the cell cycle proteins. In this study, we investigated the ability of polyphenols [Epigallocatechin Gallate (EGCG), Catechin, Catechin Gallate (CG), and Epigallocatechin (EGC)] and their degradation products [Phloroglucinol; 2,4,6 Trihydroxybenzoic acid (2,4,6 THBA); 3,4 Dihydroxyhydrocinnamic acid (3,4-DHHCA); etc.] to inhibit the CDKs (that regulate cell cycle) activity. Our pilot studies show that the different catechin compounds and their degradation products are capable of inhibiting different CDKs, suggesting their potential role in cancer prevention. We are currently investigating the mechanism of inhibition of CDKs by these compounds utilizing biochemical and molecular docking studies. Elucidation of this pathway will suggest the role of polyphenols in the prevention of CRC as well as assist in the identification of novel compounds that can be used for both cancer prevention and therapy.

Citation Format: Ranjini Sankaranarayanan, Rakesh Dachineni, Siddharth Kesharwani, Ramesh Kumar Dhandapani, Hemachand Tummala, Jayarama B. Gunaje. Identification of novel natural compounds as potential inhibitors of cyclin dependent kinases [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 4411.
Abstract 4436: Ubiquitin-specific peptidase 24 decreases c-Myc expression to inhibit lung cancer formation
Authors: Hung Jan-Jong , Chang Wen-Chang
Published: August 2019

Lung cancer is a malignant lung tumor characterized by high incidence and motility. c-Myc is a transcription factor that plays an important role in oncogenic activation to influence cellular metabolism and proliferation. Ubiquitin-specific peptidase 24 (USP24) is one of the members of USPs family to regulate protein ubiquitination. However, it is lack of evidence to uncover the role of USP24 in cancer formation. In this study, we found that cell proliferation was significantly increased by USP24 knockdown in A549 cells as accompanied by the increase of c-Myc protein. In particular, the RNA level and protein stability of c-Myc were not affected by USP24 knockdown, suggesting that USP24 affects c-Myc through regulating the translational pathway. Furthermore, c-Myc participated in USP24-knocked down-induced proliferation, not migration. To further clarify the mechanism underlying the regulation of c-Myc by USP24, we investigated whether mTOR pathway is involved in c-Myc upregulation by USP24 knockdown. Herein rapamycin, the mTOR inhibitor, prevented c-Myc upregulation caused by USP24 knockdown, suggesting that USP24 affect the protein level of c-Myc by regulating mTOR-mediated translation. We found that the downstream signaling of mTOR is activated by USP24 knockdown, including S6K and RPS6 phosphorylation. In addition, USP24 knockdown induced the binding of activated PRS6 to 5′-UTR of c-Myc, indicating that USP24 regulates c-Myc through mTOR-mediated translation. However, USP24 did not affect the phosphorylation of mTOR at Ser-2448, implying that other phosphorylation residue(s) within mTOR might be regulated by USP24. Since mTOR pathway is critical for many cancers formation, understanding the role of USP24 in regulating mTOR pathway will contribute to develop strategies to fight lung cancer.

Citation Format: Jan-Jong Hung, Wen-Chang Chang. Ubiquitin-specific peptidase 24 decreases c-Myc expression to inhibit lung cancer formation [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 4436.
Abstract 4439: MLN4924, a protein neddylation inhibitor, elicits antitumor effect via suppression of angiogenesis
Authors: Kuo Kuan-Lin , Yang Shao-Ping , Liao Shih-Ming , Pu Yeong-Shiau , Huang Kuo-How
Published: August 2019

In the process of tumorigenesis, angiogenesis provide oxygen and nutrients for tumor progression. Anti-angiogenesis has become a promising target of cancer therapy. Cullin-RING ligases (CRLs) are a cullin scaffold RING-finger domain containing E3 and are involved in the ubiquitination of specific substrates which regulate important biological processes. The holoenzyme E3 activity of CRLs is controlled by NEDD8. A novel NEDD8-activating enzyme inhibitor, MLN4924, is reported to block the neddylation of cullins and elicits antitumor effect. MLN4924 has been reported to be assoiciated with antiangiogenesis. In this study, we aim to investigate the role of anti-angiogensis in the antitumor effect of MLN4924. Our results showed that MLN4924 induces the cell cycle arrest, apoptosis and suppresses the cell viability and migration in human umbilical vein endothelial cells (HUVECs). Moreover, MLN4924 inhibits angiogenesis in Matrigel plug assay in vivo and tube-formation assay in vitro. Anti-angiogesis was involved in the antitumor effect on human urothelial carcinoma cells (T24) and colon cancer cells in vivo xenograft model. MLN4924 suppressed the VEGF-mediated VEGFR2 downstream activation.

Note: This abstract was not presented at the meeting.

Citation Format: Kuan-Lin Kuo, Shao-Ping Yang, Shih-Ming Liao, Yeong-Shiau Pu, Kuo-How Huang. MLN4924, a protein neddylation inhibitor, elicits antitumor effect via suppression of 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 4439.
Abstract 4442: CUL7 promotes cancer cell survival through promoting Caspase-8 ubiquitination
Authors: Kong Yanjie , Wang Zehua , Zhou Zhongmei , Mao Xiaoyu , Chen Ceshi
Published: August 2019

The Cullin 7 (CUL7) gene encodes a member of the cullin family of E3 ubiquitin ligases. Accumulated evidence suggests that CUL7 is oncogenic. However, the mechanism by which CUL7 improves cancer cell survival has not been fully elucidated. Here, we reported that CUL7 confers anti-apoptotic functions by interacting with Caspase-8. CUL7 prevents Caspase-8 activation by promoting Caspase-8 modification with non-degradative polyubiquitin chains at K215. CUL7 knockdown sensitized cancer cells to TRAIL-induced apoptosis in vitro and in nude mice. These results suggest that CUL7 limits extrinsic apoptotic signaling by promoting Caspase-8 ubiquitination.

Citation Format: Yanjie Kong, Zehua Wang, Zhongmei Zhou, Xiaoyu Mao, Ceshi Chen. CUL7 promotes cancer cell survival through promoting Caspase-8 ubiquitination [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 4442.