bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1291: Master regulator analysis of paragangliomas carrying SDH, VHL, or MAML3 genetic alterations
Authors: Smestad John A. , Maher Louis J.
Published: August 2018

Background: Succinate dehydrogenase (SDH) loss and mastermind-like 3 (MAML3) translocation are two clinically important genetic alterations that occur in subtypes of human paraganglioma and pheochromocytoma (PPGL) neuroendocrine tumors that correlate with increased rates of metastasis. Although hypotheses exist for how global activation of pseudohypoxia and epigenomic hypermethylation are driven via dioxygenase poisoning in the context of SDH loss, it remains unclear how these defects drive tumorigenesis. Additionally, MAML3 translocation has recently been identified as a genetic alteration in PGL and is poorly understood. We hypothesize that a key to understanding the tumorigenic basis for these genetic alterations is the deconvolution of the gene expression profiles in these tumors and identification of the transcription factors responsible for the observed transcriptional changes.

Methods: In the present work, we leverage publicly-available human tumor gene expression profiling experiments (N=179) to reconstruct a PPGL tumor-specific transcriptional network. Inferred network structure was subsequently validated by examining the frequency of known transcription factor DNA-binding motifs in the inferred regulons. We subsequently use the inferred transcriptional networks and known gene expression signatures for specific PPGL subtypes to perform a master regulator analysis nominating specific transcription factors that may control observed oncogenic patterns of gene expression. The findings of the master regulator analysis were confirmed by duplication of the analysis on a second PPGL-specific inferred transcriptional network (N=188 specimens).

Results: A small number of master regulator transcription factors are likely drivers of the observed subtype-specific gene expression patterns in SDH loss- and MAML3 translocation-positive PPGL. Unexpectedly, hypoxia-inducible transcription factors were not identified as master regulators in SDH-loss tumors. Additionally, analysis of the MAML3 translocation subtype master regulators revealed that a single transcription factor, IRX4, may control 37% of the deferentially expressed genes in this highly malignant tumor subtype.

Conclusions: These observations from unbiased analyses suggest a basis for future development of targeted therapies for specific PPGL molecular subtypes.

Citation Format: John A. Smestad, Louis J. Maher. Master regulator analysis of paragangliomas carrying SDH, VHL, or MAML3 genetic alterations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1291.
Abstract 1305: A comprehensive investigation of immune infiltrates in hepatocellular carcinoma
Authors: Yoon Sangho
Published: August 2018

Hepatocellular carcinoma (HCC), a major form of primary liver cancer, is one of the leading causes of cancer-related death, and is caused by a myriad of risk factors from chronic inflammation by viral infection to genetic defects. Recent investigations shed light on the importance and role of immune cells in tumor microenvironment; however, since a majority of studies have focused only on tumor sites, the role of immune cells in tumor-surrounding (or precancerous) area with chronic inflammation was not investigated in HCC in detail. For the treatment of HCC patients, liver transplant (total hepatectomy) has been reported to offer much longer survival and better outcome compared to patients who had local tumor resection (partial hepatectomy). Nonetheless, the significance of tumor-infiltrating immune cells (TICs) established in patients who had suffered from chronic inflammation is still vague. Here, we collected total 122 Korean HCC samples, followed by high-throughput RNA sequencing (RNA-seq), from various premalignant stages—fibrosis and dysplastic nodules, tumors to noncacerous normal liver tissues—enhanced with public RNA-seq datasets including HCC from TCGA. Decomposing immune cell types using the RNA-seq data revealed that TICs are dynamically changed with showing no or weak correlations in the normal liver tissues, while being intertwined into dense and strong correlation networks as the disease progresses. Intriguingly, patients who had total hepatectomy showed different composition of immune cells and longer disease-free survival compared to the partial hepatectomy group; however, the presence of regulatory T cells (Tregs) before the transplantation dampened this outcome in the total hepatectomy patients. Tregs in the total hepatectomy group were found to be specific to tumor sites, and were enriched upon tumor progression, while pretreatment, such as embolization, increased the presence of Tregs. Besides the specific immune cells in tumor sites, the composition of certain types of immune cells in tumor-surrounding inflamed tissues was also important for predicting patients' outcomes in HCC. Particularly, alternatively activated macrophages (M2-type) have more predictive power in the patients' outcome when they were inferred from the tumor-surrounding inflamed samples. Our study not only provides a dynamic map of tumor-surrounding and -infiltrating immune cells, but also offers understanding about immune cell networks in the progression of HCC.

Citation Format: Sangho Yoon. A comprehensive investigation of immune infiltrates in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1305.
Abstract 131: Neutrophil extracellular trap mediate mitochondrial biogenesis in cancer cells to promote growth of metastases
Authors: Yazdani Hamza , Tohme Samer , Roy Eva , Loughran Patricia , Windth Dirk Vander , Huang Hai , Tsung Allan
Published: August 2018

Neutrophils through the release of neutrophil extracellular traps (NETs) in response to surgical stress were shown to be able to capture circulating cancer cells, and in doing so, support the development of metastatic disease. In addition, NETs persist within the tumor microenvironment long after the surgical stress happens. The exact nature of the NETs-tumor interaction and the role that NETs play in dictating the tumor microenvironment is unclear. In vivo, subcutaneous and liver metastasis models of colorectal and hepatocellular cancer were performed in wild type and PAD4 KO mice that lack the ability to form NETs. In vitro, bioenergetics profiles of both HCC and CRC cancer cells were studied after co-culture with NETs, isolated from human and murine neutrophils. We found a 3-fold decrease in the tumor volume of PAD4 KO mice compared to control mice 3 weeks after tumor injection in both subcutaneous and liver metastatic models. PAD4 KO tumors showed significantly decreased neutrophil infiltration and no NET formation within the tumor compared to WT mice. PAD4 KO tumors had decreased proliferation rates and increased apoptosis. In vitro, stressed cancer cells release stress signals such as cytokines, chemokines, and DAMPs (HMGB1 and histones), that enhanced neutrophil migration towards them and subsequently triggered NET formation. In turn, NETs resulted in a 2-fold increase in the expression of mitochondrial biogenesis regulators in cancer cells. Increase mitochondrial mass was observed when treated cancer cells with NETs. A significant increase mitochondrial oxygen consumption rate (OCR) and ATP levels were observed when compared with DNase treatment. Taken together, we suggest that cancer cells help in recruiting NETs which in turn regulates mitochondrial bioenergetics for tumor growth and metastasis.

Citation Format: Hamza Yazdani, Samer Tohme, Eva Roy, Patricia Loughran, Dirk Vander Windth, Hai Huang, Allan Tsung. Neutrophil extracellular trap mediate mitochondrial biogenesis in cancer cells to promote growth of metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 131.
Abstract 1311: Integrating network biology and polypharmacology to reveal TAMS/CXCL-1 as key mediator of XIAOPI formula preventing breast cancer metastasis
Authors: Wang Neng , Wang Zhiyu , Zheng Yifeng , Wang Shengqi , Zhang Fengxue , Lin Yi
Published: August 2018

Purpose: Network pharmacology has become a powerful means of understanding the mechanisms underlying the action of Chinese herbs in cancer treatment. This study aims to validate the preventive effects and molecular mechanisms of a clinical prescription XIAOPI formula against breast cancer.

Experimental Design: The anti-metastasis ability of XIAOPI was validated by MMTV-PyMT mice and in vitro cell models. A network pharmacology-based method, which combined large-scale text-mining, drug-likeness filtering, target prediction and network analysis, was conducted to predict TAMS/CXCL-1 as key mediator of XIAOPI formula preventing breast cancer metastasis, further supported by cytokine array analysis in vitro and in vivo.

Results: In vivo breast cancer xenograft data showed that XIAOPI delayed breast cancer development and efficiently inhibited lung metastasis, accompanied by prolonged survival benefits and decreased cancer stem cell subpopulations. However, similar phenomenon were not observed in a cell model. The herb-ingredient-target network analysis further identified a total of 81 genes closely correlated with the breast cancer chemoprevention effects of XIAOPI. Cytokine array analysis further validated CXCL-1 as the key target of XIAOPI both in vitro and in vivo. Evaluation of the mechanism demonstrated that CXCL-1 administration significantly abrogated the metastatic inhibition effects of XIAOPI on breast cancer migration, invasion, stem cells subpopulations, epithelial-mesenchymal transition(EMT), or mammosphere formation abilities.

Conclusions: Overall, our study not only provides experimental evidence and molecular mechanisms that may facilitate the safe and effective use of herbal medicine for the prevention of breast cancer growth or metastasis, and may lead to CXCL-1-based therapeutic strategies for mammary malignancies.

Citation Format: Neng Wang, Zhiyu Wang, Yifeng Zheng, Shengqi Wang, Fengxue Zhang, Yi Lin. Integrating network biology and polypharmacology to reveal TAMS/CXCL-1 as key mediator of XIAOPI formula preventing breast cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1311.
Abstract 1317: Multi-scale mapping of the physical and functional architecture of the cancer cell
Authors: Zheng Fan , Yu Michael K. , Kim Minkyu , Ono Keiichiro , Flagg Mitchell , Kreisberg Jason F. , Krogan Nevan , Ideker Trey
Published: August 2018

Cancer is governed by modular systems of genes, the composition and organization of which remains poorly understood. Here, we integrate physical and functional networks from a wide range of molecular studies to assemble a comprehensive multi-scale map of human cancer cell biology. This map consists of a hierarchical catalog of protein complexes, signaling pathways and inter-pathway crosstalk implicated in cancer, and it suggests many uncharacterized functional modules as intriguing hypotheses for further validation. Analysis of the pattern of somatic mutations in The Cancer Genome Atlas (TCGA) reveals that these mutations target systems of varying scales above the level of individual genes. The map also provides a platform to integrate and interpret new 'omics data; we integrate new protein-protein interactions identified using AP-MS in multiple breast cancer cell lines, revealing how different functional modules are rewired in cancer cells. A general model browsing tool has been created to visualize and navigate these hierarchical cancer maps. This multi-scale mapping approach elucidates the molecular heterogeneity of cancer, connects tumor genotypes to phenotypes and, ultimately, enables a platform for cancer precision medicine.

Citation Format: Fan Zheng, Michael K. Yu, Minkyu Kim, Keiichiro Ono, Mitchell Flagg, Jason F. Kreisberg, Nevan Krogan, Trey Ideker. Multi-scale mapping of the physical and functional architecture of the cancer cell [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1317.
Abstract 1319: Edge-based integration of multi-omics data gives more accurate prognostic prediction in pancreatic cancer
Authors: Sun Yidi , Li Chen , Zeng Rong
Published: August 2018

Pancreatic cancer (PC) is the fourth leading cause of cancer death worldwide, and predicted to be the second within a decade. Five-year survival of PC is less than 5% with a median survival just a couple of months. Therefore, a better understanding of the molecular pathology of PC is an urgent need to achieve advances in clinical treatment for patients. Genomic analyses previously have revealed heterogeneous landscapes of mutation, copy number variation, structural variation and gene expression in pancreatic cancer. While clinical evidence for this proposition is limited and the signaling and biological effects of genomic variations are not routinely determined in human tumors even though they are rationally considered to be drug targets. Luckily, with recent advances in mass spectrometry (MS), comprehensive proteomic and lipidomics analyses provide a potentially valuable approach to validate genomic findings and discover targeted treatments for pancreatic cancer patients.

We generated high-throughput proteomic and lipidomic data for pancreatic cancer patients aged from 40 to 80 years old with matched normal samples using TMT-labeling quantitative proteomic method and target lipidomic method. We found promising protein and lipid biomarkers closely related with the clinical outcome of pancreatic cancer patients. Taking signaling and biologically pathways into consideration, we used an edge-based method to distinguish prognostics, where edge features were constructed based on the correlation between each pair of molecules. We then performed feature selection and prognostic classification for both the original expression dataset and the transformed edge dataset and found the edge-based features can give a more accuracy for prognostic prediction, suggesting the promising role of protein and lipid edge biomarkers for clinical utility in pancreatic cancer.

Our preliminary results highlight the usefulness of edge-based integration of proteomics and lipidomics data. To explain the mechanisms in the development of pancreatic cancer, we are going to integrate genomic variations from whole genome sequencing (WGS) into our current progress for further analysis. In conclusion, this initial work demonstrated a strategy that may enable more accurate prediction of the survival of pancreatic cancer by integrating multi-omics data from genomics, proteomics and lipidomics. In principle, we anticipate the declaration of possible mechanisms enabling the treatment of pancreatic cancer patients in the near future.

Citation Format: Yidi Sun, Chen Li, Rong Zeng. Edge-based integration of multi-omics data gives more accurate prognostic prediction in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1319.
Abstract 1321: Mutational landscape of adherens junctions in skin cutaneous melanoma
Authors: Korla Praveen Kumar , Gracilla Daniel Esguerra , Kuo Brian Yu-Ting , Sheu Jim Jinn-Chyuan
Published: August 2018

Cadherin proteins regulate cell-cell contact through calcium-dependent manner, thus play pivotal roles in morphogenetic and differentiation processes during tissue development. Reprograming in cadherin profiles contributes to epithelial-mesenchymal transition (EMT), leading to disorganized lining of epithelial tissues and cancer metastasis. To elucidate genetic impacts of classical cadherins on melanoma development, mutation profiles were analyzed using data from the TCGA databank. Among 20 classical cadherins, CDH6 showed the highest mutation frequency (11.3 %), and most of which were G to A or C to T nucleotide substitutions, a signature for UV-caused DNA damage. Two groups of unique hotspots were found on the fifth cadherin domain (D5) (S524L and S526F) and cytoplasmic cadherin domain (CDH-C) (E662K and E722K), respectively. Protein interactome and expression co-occurrence analyses placed CDH6 at the center of core cadherin network and showed strong heterophilic interactions of CDH6 with CDH4, CDH9, and CDH12. Multiple sequence alignment indicated the involvement of those hotspots in the conserved regions, which are responsible for heterophilic interaction on D5 or catenin (alpha, beta, and gamma) binding on CDH-C. Molecular modeling defined alterations in protein structures of the D5 and CDH-C domains caused by the hotspot mutations, which may influence cell-cell contact and the down-stream signaling. In sum, our data suggest CDH6 as the key player to modulate adherens junctions during the development of melanoma.

Citation Format: Praveen Kumar Korla, Daniel Esguerra Gracilla, Brian Yu-Ting Kuo, Jim Jinn-Chyuan Sheu. Mutational landscape of adherens junctions in skin cutaneous melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1321.
Abstract 1336: Autophagy inhibition reverses EMT and reduces 3D growth of pancreatic adenocarcinoma cells in vitro and in vivo
Authors: Pearce Jane B. , O'Flanagan Ciara H. , Hursting Stephen D.
Published: August 2018

Pancreatic cancer is the 4th most fatal cancer in US men and women, with 5-year survival rates under 5%. Obesity is one risk factor known to increase pancreatic cancer risk by facilitating tumorigenesis and metabolic processes associated with pancreatic tumor growth and progression. Autophagy is the catabolic process of recycling cellular organelles to provide energy to the cell and is often induced when cells become stressed. Autophagy plays a critical role in cancer growth and metastasis, and in many cancers autophagy inhibition decreases the size of tumors in in vivo models. Our current studies investigate the link between the autophagy inhibition and the invasiveness of pancreatic cancer. To understand the role of autophagy on invasion, EMT and 3D growth in pancreatic cancer, Kras-driven Panc02 mouse pancreatic cancer cells were transduced using CRISPR/Cas9 to cleave the gene encoding autophagy related protein 5 (Atg5). Atg5 deletion resulted in striking morphological alterations, including a more epithelial-like appearance. In addition, Atg5 deletion resulted in increased expression of the epithelial marker E-Cadherin, concurrent with decreased expression of mesenchymal markers N-Cadherin and Snail. Furthermore, Atg5 deletion resulted in significant decreased invasiveness and ability to form colonies under anchorage-independent conditions, with no significant difference in 2D cell growth. Orthotopic, intrapancreatic injection of these cells into C57BL/6 mice resulted in reduced tumor take and growth. Taken together, these results for the first time show that autophagy inhibition results in reduced EMT and 3D growth in pancreatic adenocarcinoma and highlight autophagy's role in pancreatic adenocarcinoma progression.

Citation Format: Jane B. Pearce, Ciara H. O'Flanagan, Stephen D. Hursting. Autophagy inhibition reverses EMT and reduces 3D growth of pancreatic adenocarcinoma cells in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1336.
Abstract 134: Dysbiosis of gut microbiota in chronic stressed patients with endometriosis
Authors: Xu Jing , Kang Yu , Yu Yinhua
Published: August 2018

Although endometriosis is a benign disease, it has malignant characters such as invasion and migration. Besides, there is an endometrial type of ovarian cancer, which means endometriosis is similar to malignant disease at some extent. Existing researches showed that patients with endometriosis often have high level of chronic stress. Furthermore, chronic stress can promote the growth of endometriosis by activating the β-adrenergic signaling pathway. Gut microbiota is reported to be related to the stress level through the gut-brain axis. So we wonder whether gut microbiota is associated with chronic stress level in endometriosis patients, which may provide a new clinical intervention method to improve prognosis. We grouped the endometriosis patients into stress group and control group with GAD-7 and PHQ-9 questionnaires and collected patients' fecal and tissues samples. Gut microbiota compositions were analyzed by the 16SrRNA gene sequencing-based method and we performed Immunohistochemistry (IHC) to detect the activation of inflammatory pathways in endometriosis tissues. We found that in patients with endometriosis, dysbiosis of gut microbiota is associated with patients' stress levels and Paraprevotella, Odoribacter, Veillonell and Ruminococcus show significant differences with reduced levels in chronic stressed endometriosis patients group, which represents a disease-specific change of gut microbiota on the genus level. Besides, compared to the control group, the expression level of inflammatory cytokines, NF-κB p65 and COX-2, increased in the chronic stressed patients group, which means that dysbiosis of gut microbiota may active the inflammatory pathway of gut-brain axis. In conclusion, we found new disease-specific changes of gut microbiota in chronic stressed patients with endometriosis, which can be a new examination target of chronic stress level and may provide an adjuvant therapy method for clinical psychological intervention, reducing the stress level and improving the prognosis of endometriosis patients.

Citation Format: Jing Xu, Yu Kang, Yinhua Yu. Dysbiosis of gut microbiota in chronic stressed patients with endometriosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 134.
Abstract 1340: Assessing autophagic flux in cell culture models with a novel plate-based assay
Authors: Lazar Dan F. , Kessens Ryan W. , Gillette Amani A. , Butler Braeden L. , Eggers Christopher T. , Binkowski Brock F. , Vidugiris Gediminas , Slater Michael R. , Ma Dongping , Cali James J.
Published: August 2018

The importance of autophagy in normal cell health and diseased states, including cancer, immunology and inflammation, has become quite clear. Given the growing interest in screening for autophagy modulators, we have developed a homogeneous plate-based assay to measure autophagic flux in cell culture models using NanoLuc® Binary Technology (NanoBiT™). LC3B (Atg8) protein was tagged on its N-terminus with an 11 amino acid peptide, (HiBiT), as well as an intervening “spacer” sequence. When expressed at low-to-moderate levels in mammalian cell lines, this LC3-based reporter is subject to degradation by the autophagic pathway. Changes in reporter levels following cell treatment are determined by addition of a lytic reagent containing Large BiT (LgBiT), which rapidly associates with HiBiT to produce a bright, luciferase activity in the presence of added substrate. In stable U2OS reporter cells, mTORC inhibitor treatment decreased assay signal by 25% (rapamycin) to 60% (PP242, AZD8055), consistent with compound stimulation of autophagic flux and degradation of the autophagy reporter. In contrast, treatment with autophagy inhibitors (bafilomycin A1, chloroquine) produced a 70-80% increase in assay signal, consistent with accumulation of autophagy reporter following blockade of basal autophagy. Similar results were obtained in HEK293 cells stably expressing the reporter, as well as in cells in which autophagy reporter was transiently expressed via simple, one-step transduction with BacMam viral particles. Mechanism of action of autophagy modulators was confirmed through blockade of effects by 50nM bafilomycin A1 cotreatment. Multiplex with a cell necrosis detection agent allowed for same-well determination of cytotoxic effects that might undermine analysis of a compound's discrete effects on autophagic flux. When assayed in 384-well plates with automation, U2OS and HEK293 autophagy reporter cells produced Z' values of 0.6-0.7 in response to autophagy induction (PP242), and Z' values of 0.7-0.8 by subsequent blockade of autophagy (PP242 + bafilomycin A1). Therefore, using this novel plate-based assay system for the determination of autophagic flux, it is possible to screen test agents and quantitatively assess their effects in cell culture models.

Citation Format: Dan F. Lazar, Ryan W. Kessens, Amani A. Gillette, Braeden L. Butler, Christopher T. Eggers, Brock F. Binkowski, Gediminas Vidugiris, Michael R. Slater, Dongping Ma, James J. Cali. Assessing autophagic flux in cell culture models with a novel plate-based assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1340.
Abstract 1346: Lycorine Promotes Autophagy and Apoptosis via TCRP1/Akt/mTOR Axis Inactivation in Human Hepatocellular Carcinoma
Authors: Yuhaiyang Yuhaiyang Yuhaiyang
Published: August 2018

Lycorine is a multi-functional bioactive compound, and it possesses potential anti-cancer activities. However, little is known about the underlying mechanism. In this research we have found that lycorine significantly induces the apoptotic and autophagic capacities of hepatocellular carcinoma (HCC) cells in vitro and in vivo. Treatment with specific autophagy inhibitor (3-methyladenine/Bafilomycin A1) or knockdown of LC-3B/Atg5 by siRNA drastically enhances the apoptotic cell death effect by facilitating the switch from autophagy to apoptosis. Molecular validation mechanistically demonstrates that lycorine-induced apoptosis and autophagy in HCC cells is associated with decreased protein levels of tongue cancer resistance-associated protein 1 (TCRP1), and we further find that inhibition of TCRP1 decreases phosphorylation level of Akt and represses Akt/mTOR signaling. Finally, lycorine-induced apoptosis and autophagy suppress the growth of xenograft hepatocellular tumors without remarkable toxicity. Our results elucidate a novel molecular mechanism whereby lycorine promotes apoptosis and autophagy through TCRP1/Akt/mTOR pathway in HCC. Our results reveal that lycorine might be a potential therapeutic agent for the treatment of HCC.

Citation Format: Yuhaiyang Yuhaiyang Yuhaiyang. Lycorine Promotes Autophagy and Apoptosis via TCRP1/Akt/mTOR Axis Inactivation in Human Hepatocellular Carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1346.
Abstract 1349: Functional and mechanistic characterization of PRMT6 regulated autophagy in hepatocellular carcinoma
Authors: Che Noélia , Tong Man , Ng Kai Yu , Kau Phillis WF , Guan Xin Yuan , Huen Michael SY , Ma Stephanie
Published: August 2018

Hepatocellular carcinoma (HCC) is a major health burden worldwide. Tumor recurrence and therapy resistance represent major obstacles in the treatment of the disease, with their mechanism of action largely uncharacterized. Autophagy is a critical survival factor for cancer cells, whereby it maintains cellular homeostasis including degradation of damaged organelles and unwanted proteins as well as the support of cellular biosynthesis in response to environmental stress, preventing cells from undergoing apoptosis. Our group has recently reported on the regulatory role of protein arginine methyltransferase 6 (PRMT6) down-regulation in maintenance of HCC, particularly potentiating resistance to sorafenib and chemotherapy. By tandem affinity purification/mass spectrometry analysis of cells with PRMT6 stably overexpressed, we identified a number of autophagy related proteins as potential interacting partners of PRMT6, suggesting a possible role of PRMT6 in modulating autophagy in HCC. Treatment of HCC cells with stress induced conditions, including sorafenib, nutrient deprivation and hypoxic condition, resulted in a marked reduction of PRMT6 expression, concomitant with elevated levels of LC3BII. Further, under these various stimuli, knockdown of PRMT6 in HCC cells would also result in enhanced expression of LC3BII, accumulation of GFP-LC3B positive puncta staining as well as an increase in autophagosome formation. Consistently, treatment of HCC cells with PRMT6 overexpressed would leading to an opposing effect. Further studies by immunoprecipitation analysis validated PRMT6 to physically interact with AMBRA1 (activating molecule in BECN1-regulated autophagy protein 1), a well-known autophagy initiative component, which takes part in autophagy vesicle nucleation, a critical step for autophagic initiation. Our findings suggest PRMT6 down-regulation in HCC tumors to promote tumorigenicity through autophagic flux de-regulation. Targeting the mechanisms of stress response may provide novel therapeutic insights for the disease.

Citation Format: Noélia Che, Man Tong, Kai Yu Ng, Phillis WF Kau, Xin Yuan Guan, Michael SY Huen, Stephanie Ma. Functional and mechanistic characterization of PRMT6 regulated autophagy in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1349.
Abstract 1351: Inhibition of autophagy enhances dihydroartemisinin (DHA)-induced cytotoxicity through DR5 upregulation
Authors: Mei-Chuan Chen
Published: August 2018

Artemisinin (Qinghaosu) comes originally from Qinghao has been used against Plasmodium falciparum malaria for a long time. Dihydroartemisinin, an analogue of artemisinin, is also be listed by WHO to exhibit antimalarial and anticancer properties.

In our study, DHA exhibits antiproliferative activity in several solid tumors, especially against colorectal cancer (CRC) HCT116 cells. Our current results show DHA suppresses cell proliferation and induces caspase-dependent apoptosis without inducing necrosis-like cell death in HCT116 cells. In addition, DHA interferes autophagy pathway by upregulating of LC3 II formation in HCT116 cells. Inhibition of autophagy with either autophagy inhibitor 3-methyladenine (3MA) or KD of ATG5 potentiated DHA-induced cell death. We found DHA increased expression level of DR5 while had no effect on DR4 expression level on surface of tumor cells. Further, cell-surface DR5 expression was significantly elevated in DHA-treated ATG5-KD cells. These findings suggest that inhibition of the autophagy enhances DHA-induced cytotoxicity through DR5 upregulation.

Key words: Apooptsis, DR5 (Death Receptor 5), autophagy, colorectal cancer

Citation Format: Chen Mei-Chuan. Inhibition of autophagy enhances dihydroartemisinin (DHA)-induced cytotoxicity through DR5 upregulation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1351.
Abstract 1363: Regulation of BRCA1 by SIRT2
Authors: Minten Elizabeth , Zhang Hui , Li Chunyang , Head PamelaSara , Yu David
Published: August 2018

In our everyday lives, we are constantly being exposed to DNA damaging agents, both from internal and external sources. The most deleterious type of DNA damage a cell can suffer is the DNA double-stranded break (DSB). Breast Cancer 1 (BRCA1) is a protein necessary for the proper repair of resultant DNA DSBs through the homologous recombination (HR) pathway. Defects in BRCA1 have been linked to different types of cancer in both men and women, including breast, ovarian, and pancreatic cancer. However, the regulation of BRCA1 is not yet well understood, and in many cases, how defects in this pathway lead to an increased risk of developing cancer is also unknown, making preventative care and treatment of resulting cancers more difficult. Our lab has discovered that SIRT2, a histone deacetylase and putative human tumor suppressor, plays a crucial role in the DNA damage response (DDR) and repair of DNA DSBs. We have shown that depletion of SIRT2 impairs HR and increases cell sensitivity to IR in a deacetylase-dependent manner. A mass spectrometry analysis showed SIRT2 interacts with a number of proteins involved in DDR, including BRCA1. We validated the interaction between SIRT2 and BRCA1 and also found SIRT2 deacetylates BRCA1 both in vitro and in cells. Depletion of SIRT2 and subsequent deacetylation of BRCA1 decreases BRCA1 protein levels in cells and thus impairs HR. Our results show SIRT2 is a novel regulator of BRCA1 and is critical for the repair of DNA DSBs through HR. These findings provide insight into how dysregulation of SIRT2 or BRCA1 can result in genomic instability and lead to the development of cancer.

Citation Format: Elizabeth Minten, Hui Zhang, Chunyang Li, PamelaSara Head, David Yu. Regulation of BRCA1 by SIRT2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1363.
Abstract 1366: MBD4 guards against DNA damage from methylcytosine deamination
Authors: Chew Edward , Sanders Mathijs A. , Flensburg Christoffer , Zeilemaker Annelieke , Miller Sarah E. , Hinai Adil S. al , Bajel Ashish , Luiken Bram , Rijken Melissa , Mclennan Tamara , Hoogenboezem Remco M. , Kavelaars François G. , Blewitt Marnie E. , Bindels Eric M. , Alexander Warren S. , Löwenberg Bob , Roberts Andrew W. , Majewski Ian J. , Valk Peter J.
Published: August 2018

Spontaneous methylcytosine (5mC) deamination is a common source of cytosine to thymine (C>T) mutations. These mutations accumulate over time, serving as a “molecular clock” that tracks cellular age. MBD4 is a thymine glycosylase that recognises sites of 5mC deamination and initiates base excision repair.

Three patients (of whom 2 are siblings) with germline MBD4 loss of function (LOF) mutations developed acute myeloid leukemia (AML) at a young age (<35 years old). Whole exome (WES) and whole genome sequencing (WGS) showed an elevated C>T mutation rate ~33 times the average for AML. The mutations occurred nearly exclusively at CG dinucleotides. Reduced representation bisulfite sequencing showed the mutations occurred at previously methylated cytosines. The 3 AMLs had somatic C>T mutations in the same driver genes (DNMT3A and either IDH1 or IDH2), suggesting the methylation damage charts a recurrent path towards malignancy in the hematopoietic system.

MBD4 is inactivated in other cancers, although this appears to be a rare event. Nine of 10,638 cancers in the TCGA database had MBD4 LOF mutations. In a uveal melanoma and a glioblastoma multiforme, there was also loss of heterozygosity of the wild-type MBD4. These 2 cancers with MBD4 deficiency exhibit the same mutational signature, with a high C>T mutation rate at CG dinucleotides.

To verify the link between MBD4 and the 5mC damage signature, Mbd4-/- and Mbd4+/+ mouse bone marrow were cultured in semi-solid agar and WGS was performed on individual myeloid progenitor colonies. Mbd4-/- myeloid progenitor colonies displayed the same increase in C>T mutations at CG dinucleotides. This highlights the importance of MBD4 across species.

Mbd4 deficient mice offer a model system to investigate mutation acquisition and cancer pathogenesis. Work on the interaction of MBD4 with other leukaemia initiating genes is underway.

Citation Format: Edward Chew, Mathijs A. Sanders, Christoffer Flensburg, Annelieke Zeilemaker, Sarah E. Miller, Adil S. al Hinai, Ashish Bajel, Bram Luiken, Melissa Rijken, Tamara Mclennan, Remco M. Hoogenboezem, François G. Kavelaars, Marnie E. Blewitt, Eric M. Bindels, Warren S. Alexander, Bob Löwenberg, Andrew W. Roberts, Ian J. Majewski, Peter J. Valk. MBD4 guards against DNA damage from methylcytosine deamination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1366.
Abstract 1367: BRCA1 and BARD1 protein interactions that are required for DNA repair function
Authors: Adamovich Aleksandra , Wingo Margaret , Banerjee Tapahsama , Gardner Miranda , Freitas Michael , Parvin Jeffrey
Published: August 2018

Breast and ovarian cancers are prevalent among women, and hereditary breast and ovarian cancers (HBOCs) have been associated with germline mutations in genes such as BRCA1 and BARD1. BRCA1 and BARD1 form an obligate heterodimer, and the BRCA1/BARD1 complex is required for tumor suppression functions. Our lab has tested hundreds of BARD1 and BRCA1 variants and identified many that are deficient in homologous recombination, which we have shown to accurately predict cancer predisposition in the clinic. Several of these functionally defective BRCA1 mutants map to a pocket of amino acids on the surface of the protein that does not have any known binding partners. Repair-deficient mutations are also present on the surface of the BARD1 protein in domains that are not known to be associated with DNA repair. We hypothesize that the DNA repair deficiencies mediated by these BRCA1 and BARD1 mutants are due to differences in protein binding when compared to wild-type protein. We have found that BRCA1 mutants in this protein pocket do not phosphorylate and do not localize to the nucleus following DNA damage, both of which are characteristic of the DNA damage response. However, BARD1 repair-deficient mutants still bind phosphorylated BRCA1, indicating that their deficiencies are not due to loss of BRCA1 function. To investigate protein interaction differences we are creating fusion proteins of wild-type and mutant BRCA1 with BioID2, which will biotinylate proteins that bind to BRCA1. We will then be able to identify, via avidin purification and mass spectrometry, protein interactions that are present in the wild-type but absent in the mutant BRCA1. Novel proteins that we identify will be tested for DNA repair and tumor suppressor function. This information will allow us to better understand both BARD1 and BRCA1 function and the mechanism of homologous recombination.

Citation Format: Aleksandra Adamovich, Margaret Wingo, Tapahsama Banerjee, Miranda Gardner, Michael Freitas, Jeffrey Parvin. BRCA1 and BARD1 protein interactions that are required for DNA repair function [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1367.
Abstract 1373: Circular RNAs generated from the BRCA1 pseudogene regulates the DNA damage response through SERBP1 RNA-binding protein
Authors: Han Yoo J. , Zhang Jing , Mason Jennifer M. , Yoshimatsu Toshio F. , Du Xinxin , Hurley Ian , Kim Danny E. , Anantharaman Aparna , Brunet Laia P. , Prat Aleix , Kwon John , Prasanth Kannanganattu V. , Olopade Olufunmilayo I.
Published: August 2018

The genomic region encompassing the BRCA1 gene includes the BRCA1P1 pseudogene (BRCA1P1) within ~170kb at chromosome 17q21. The homology and proximity of BRCA1 and BRCA1P1 create a hot spot for recombination, resulting in a large genomic rearrangement found in some families with breast and ovarian cancers. While the significant role of BRCA1 in regulating the DNA damage response (DDR) is well defined, little is known about the biological properties of the BRCA1P1 pseudogene. To functionally annotate BRCA1P1, we conducted a CRISPR-Cas 9 knockout of the pseudogene, as well as knockdown of the expression using antisense oligonucleotides. Both knockout and knockdown of the pseudogene result in increases in apoptosis and sensitivity to DNA damaging drugs. Depletion of BRCA1P1 accumulates spontaneous DNA damage foci and leads to replication fork stalling. Mechanical studies reveal that the BRCA1P1 generates circular RNAs, which are retained in nuclei. Mass spectrometry followed by chromatin isolation by RNA purification (ChIRP) identifies a specific association of the circular RNAs with SERBP1 RNA-binding protein, which was confirmed by ChIRP-Western. Consequently, BRCA1P1 knockout decreased SERBP1 protein expression in the nuclei. Based on the previous report that SERBP1 binds to mRNAs of DDR proteins and controls their translation, we propose a mechanism of BRCA1P1-driven regulation of DDR protein expression through SERBP1. Our data indicate that the BRCA1P1 pseudogene plays a role in regulating replication fork progression and genomic instability in breast cancer cells through an interaction with SERBP1 RNA-binding protein.

Citation Format: Yoo J. Han, Jing Zhang, Jennifer M. Mason, Toshio F. Yoshimatsu, Xinxin Du, Ian Hurley, Danny E. Kim, Aparna Anantharaman, Laia P. Brunet, Aleix Prat, John Kwon, Kannanganattu V. Prasanth, Olufunmilayo I. Olopade. Circular RNAs generated from the BRCA1 pseudogene regulates the DNA damage response through SERBP1 RNA-binding protein [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1373.
Abstract 1381: Targeting histone acetyltransferases to reprogram high C-MYC expressing cancers
Authors: Costa Elodie M. Da , Armaos Gregory , Beaudry Annie , Richer Chantal , Caron Maxime , St-Onge Pascal , Johnson Jeffrey , Krogan Nevan , Sai Yuka , Downey Michael , Sinnett Daniel , McGraw Serge , Raynal Noël J.
Published: August 2018

In cancer, epigenetic modifications are strongly altered and are responsible for gene expression aberrations. In a drug screening initiative, we recently reported that proscillaridin, a cardiac glycoside (CG), exhibits unsuspected epigenetic and anticancer activities. To understand CG's epigenetic mechanisms of action, we performed RNA sequencing analysis, which showed proscillaridin effects on global gene expressions in acute lymphoblastic leukemia cells (MOLT-4). Genes associated with apoptosis and cell differentiation were upregulated whereas master transcription factors and oncogenic pathway genes were downregulated. Mechanistic studies revealed that proscillaridin decreased histone 3 acetylation, which correlated with histone acetyltransferase (KATs) downregulation (CBP, P300, TIP60, GCN5 and MOZ). Acetylome studies by mass spectrometry showed an acetylation loss in chromatin regulators, the oncogene C-MYC and its associated proteins. Proscillaridin induced C-MYC transcript and protein degradation. Moreover, in a panel of cancer cell lines, we measured that cancer cells sensitivity to proscillaridin treatment was positively correlated with C-MYC protein levels. Conversely, proscillaridin did not affect C-MYC protein level in low C-MYC expressing cancer cell lines. For the first time, we showed that CGs target histone acetyltransferases and C-MYC oncogene in high C-MYC expressing cancers. We propose that CGs can be repurposed as new epigenetic drugs in high C-MYC expressing cancers.

Citation Format: Elodie M. Da Costa, Gregory Armaos, Annie Beaudry, Chantal Richer, Maxime Caron, Pascal St-Onge, Jeffrey Johnson, Nevan Krogan, Yuka Sai, Michael Downey, Daniel Sinnett, Serge McGraw, Noël J. Raynal. Targeting histone acetyltransferases to reprogram high C-MYC expressing cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1381.
Abstract 1386: Safety and tolerability of guadecitabine (SGI-110) plus cisplatin in patients with platinum refractory germ cell tumors (GCT): A phase 1 study
Authors: Albany Costantine , Hashemi Neda S. , Fang Fang , Lowder James , Einhorn Lawrence , Nephew Kenneth
Published: August 2018

BACKGROUND: Guadecitabine (SGI-110) is a novel hypomethylating dinucleotide of decitabine and deoxyguanosine resistant to degradation by cytidine deaminase. Germ cell tumor including platinum-resistant embryonal carcinoma cells lines are extremely sensitive to hypomethylating agents and low dose decitabine can restore cisplatin sensitivity in cell lines. We aimed to assess the safety and clinical activity of guadecitabine in combination with cisplatin in patients with platinum-refractory GCT.

METHODS: In this open-label, phase 1 study, patients with GCT refractory to or had relapsed after platinum-based treatment were treated with subcutaneous (SQ) guadecitabine, once-daily for 5 consecutive days, followed by cisplatin on day 8 in a 28-day treatment cycle. A modified toxicity probability interval (mTPI) dose-escalation design was used in which we treated patients with guadecitabine doses of 30-45 mg/m2 plus cisplatin 100 mg/m2 up to 6 cycles until progression or intolerable toxicity. The primary objective was to assess safety and tolerability of guadecitabine in combination with cisplatin, determine the maximum tolerated and identify the recommended phase 2 dose of guadecitabine. Safety analyses included all patients who received at least one dose of guadecitabine. Pharmacodynamic analyses to determine the biologically effective dose included all patients for whom samples were available. This study is registered with ClinicalTrials.gov, number NCT02429466.

FINDINGS: Between 2015 and 2017, we enrolled and treated 10 patients. Grade 3 or 4 adverse events were febrile neutropenia, thrombocytopenia, anemia, and diarrhea. The most common serious adverse events were febrile neutropenia and thrombocytopenia. Guadecitabine 45 mg/m2 was associated with prolonged SAE and felt to be unsafe to continue. The maximum tolerated dose was 30 mg/m2 daily × 5. Two patients achieved a complete response to treatment lasting more than 6 months.

INTERPRETATION: Guadecitabine SQ at 30 mg/m2 daily × 5 is well tolerated and is clinically and biologically active in patients with platinum-refractory GCT. The study is still ongoing.

Citation Format: Costantine Albany, Neda S. Hashemi, Fang Fang, James Lowder, Lawrence Einhorn, Kenneth Nephew. Safety and tolerability of guadecitabine (SGI-110) plus cisplatin in patients with platinum refractory germ cell tumors (GCT): A phase 1 study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1386.