bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1284: How can you interpret gene lists from -omics experiments
Authors: Wong Jeffrey V. , Luna Augustin , Demir Emek , Rodchenkov Igor , Babur Özgün , Sander Chris , Bader Gary D.
Published: August 2018

Understanding the mechanisms responsible for a cellular behaviour often begins with observations of genes and gene products. Depending on the type of experiment, the number of resulting genes can be small, but increasingly, researchers are faced with many thousands of measurements, as in the case of transcriptomic or protein-DNA binding observations. Here, we describe ways to pair experimental results consisting of one or more genes with analysis tools with the overall aim being to make results more biologically interpretable. In certain cases, experimental approaches such as screens for essential genes can generate one or a few ‘genes of interest' and there is a desire to understand their relationship to one another as well as discover links to additional, interesting genes. To this end, ‘GeneMANIA' is a web tool that accepts gene names and returns a network visualization of related genes based on similarity in expression, localization, protein domains and those involved in physical interactions. Likewise, ‘PCViz' is a web tool that displays a network of interactions drawn from Pathway Commons, a web resource for pathway and interaction knowledge. In cases where experiments generate a lengthy list of genes, for instance, transcriptomic measurements, there is a desire to understand their relevance to a phenotype of interest. Pathway enrichment analysis methods aim to summarize gene lists as pathways, which have a closer link to cell function. An online ‘Guide' by Pathway Commons includes workflows that illustrate how to chain together software tools to identify pathways from the corresponding gene-level data then organize and summarize the pathway-level results in an interactive visualization known as an Enrichment Map. For those wishing to drill-down to individual pathways, Pathway Commons offers a set of web apps, including ‘Search' that enables users to query by keyword and visualize ranked search results. Ongoing development of web apps aims to enhance the accessibility to pathways and integrate support for analysis and visualization of experimental data. The full complement of data, tools and resources offered by Pathway Commons in support of pathway analysis are described.

Citation Format: Jeffrey V. Wong, Augustin Luna, Emek Demir, Igor Rodchenkov, Özgün Babur, Chris Sander, Gary D. Bader. How can you interpret gene lists from -omics experiments [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 1284.
Abstract 1294: Preoperative prediction of microvascular invasion in HCC using radiomics on multisequence gadoxetic acid-enhanced MR images
Authors: Wei Jingwei , Gu Dongsheng , Dong Di , Zhang Shuaitong , Jin Yushen , Tian Jie
Published: August 2018

Micro-vascular invasion (MVI) is underscored as a judicious risk factor for early-stage recurrence and poor prognosis of hepatocellular carcinoma (HCC). Preoperative knowledge of MVI would assist in therapy decision making beforehand, thus prolong the survival of HCC patients. However, current diagnosis of MVI could only be obtained by pathological confirmation after the surgery or through punctures, which hinders timely therapeutic regime plan or causes needless suffering for the patients. In this study, we aimed to perform a preoperative prediction on MVI using multi-sequence Radiomics on gadoxetic acid-enhanced MR images, along with multi-variable clinical factor analysis. A cohort of 208 patients diagnosed with HCC was enrolled with institutional review board approval. Radiomic features including shape and size, intensity, textural and wavelet features were extracted on segmented region of interest. The least absolute regression modeling was used to select the most effective radiomic features which had potential for MVI identification. We finally utilized logistic regression modelling to generate the single radiomics signature for each sequence with Akaike information criteria as the stopping rule. Gadoxetic acid-enhanced hepatobiliary phase (HBP) T1-weighted image and HBP T1 map turned out to be the best-performed sequences with AUC over 0.7 on both training and validation cohorts. The fusion radiomics signature incorporating effective features from these two sequences achieved the optimal prediction with AUC of 0.895 on the training cohort and 0.837 on the validation cohort. Adding the pertinent clinical and radiological factors: AFP, irregular tumor border and arterial peritumoral enhancement, the final combined model could successfully predict the MVI with AUC of 0.943 on the training cohort and 0.861 on the validation cohort. Our study revealed the archetypal radiomic features related MVI and highlighted the fusion radiomics signature as a powerful imaging marker for MVI preoperative prediction. Radiomics combined with traditional clinic-radiological information would no doubt improve the clinical decision making in HCC therapy.

Citation Format: Jingwei Wei, Dongsheng Gu, Di Dong, Shuaitong Zhang, Yushen Jin, Jie Tian. Preoperative prediction of microvascular invasion in HCC using radiomics on multisequence gadoxetic acid-enhanced MR images [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 1294.
Abstract 1295: Single cell RNA sequencing of lung adenocarcinoma reveals heterogeneity of immune response-related genes
Authors: Ma Keyue , Schonnesen Alexandra A. , Eckhardt S. Gail , Jiang Ning
Published: August 2018

Immunotherapies have been the most promising approaches to treating cancers, while the partial responses in multiple clinical trials suggest the significance of characterizing intertumoral and intratumoral heterogeneity for better prognosis and decision-making in treatment. The lack of intratumoral characterization of immune response-related genes in cancer cells, however, hinders the further development of metrics to select and predict immunotherapies. Therefore, we applied single cell RNA-seq data from lung adenocarcinoma patients to identify intratumoral heterogeneity of immune response-related genes and demonstrated its potential effect on immunotherapy efficacy. We found IFNγ signaling pathway genes are heterogeneously expressed and co-regulated in cancer single cells, including MHC class II molecules (MHCII), a set of favorable prognosis markers. The upregulation of MHCII is also mutually exclusive with upregulation of cell cycle pathways, which are associated with unfavorable prognosis. Moreover, analysis of two candidates of cancer vaccination approaches, neoantigens and cancer testis antigens, revealed their ectopic expression in single cells. These analyses provide the rationale of applying combinatorial therapies to prevent tumor escape and basis for future development of prognosis metrics based on intratumoral heterogeneity.

Citation Format: Keyue Ma, Alexandra A. Schonnesen, S. Gail Eckhardt, Ning Jiang. Single cell RNA sequencing of lung adenocarcinoma reveals heterogeneity of immune response-related genes [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 1295.
Abstract 130: Cancer teammate survives longer: Role of cancer-associated a2 V-ATPase in regulation of neutrophil survival
Authors: Ibrahim Safaa A. , Kulshrestha Arpita , Katara Gajendra K. , Sahoo Manoranjan , Beaman Kenneth D.
Published: August 2018

Tumor associated neutrophils (TAN) represent a significant portion of tumor leukocytes and accumulate in many types of cancer. TAN can survive for weeks in cancer tissues and have been proposed as key mediator of malignant transformation, tumor progression, angiogenesis and in the modulation of the antitumor immunity. Identifying cancer associated factors that promote the survival and protumorigenic properties of neutrophils will provide new targets for cancer therapy. Cancer cells secrete a peptide from the Nterminal domain of a2isoform vacuolar ATPase (a2NTD) that promotes the protumorigenic properties of neutrophils that enhances angiogenesis and tumor cell invasiveness. The present study attempts to decipher the role of a2NTD in the regulation of neutrophil apoptosis. The treatment of human neutrophils with recombinant a2NTD led to 2.1 fold increase in the percentage survival of neutrophils (P < 0.001). a2NTD treatment activates NF- kBp65 by activating reactive oxygen species (ROS) generation in neutrophils. Activation of NF- kB pathway leads to upregulation of the gene expression of anti-apoptotic factors; Bcl2-A1, Bcl-xL, c-Flip, G-CSF and downregulates the pro-apoptotic factors such as BAX. Parthenolide; NF-kB inhibitor, negated the effect of a2NTD on enhancing neutrophil survival confirming the involvement of NF-kB pathway in the a2NTD mediated increase of neutrophil survival. Further analysis showed that a2NTD treatment stabilizes the neutrophil mitochondrial membrane potential and decreases Bax translocation to the mitochondria that was associated with a decrease in caspase 9 and caspase 3 activity. Since there is a direct link between the intrinsic and the extrinsic pathways of apoptosis, our results showed that there was no change in the Fas and Fas ligand expression upon a2NTD treatment. Further analysis would uncover the effect of a2NTD treatment on the extrinsic pathway of apoptosis in neutrophils. Together, our results demonstrate a novel role of the cancer associated a2-vacuolar ATPase on retarding the mitochondrial death pathway in neutrophils by the action of a2NTD.

Citation Format: Safaa A. Ibrahim, Arpita Kulshrestha, Gajendra K. Katara, Manoranjan Sahoo, Kenneth D. Beaman. Cancer teammate survives longer: Role of cancer-associated a2 V-ATPase in regulation of neutrophil survival [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 130.
Abstract 1306: Density-based mutation clustering in 3D space
Authors: Weerasinghe Amila , Sengupta Sohini , Scott Adam D. , Bailey Maththew H. , Wendl Michael C. , Chen Ken , Mills Gordon , Ding Li
Published: August 2018

Cancer driver mutation and gene discovery has been a major challenge in cancer research. Several computational tools focus on clustering mutations on 3D protein structures to identify cancer drivers, including HotSpot3D, CLUMPS, HotMAPS, 3DHotSpots.org and e-Driver. While other tools provide a single snapshot of mutation clusters on the 3D structure, HotSpot3D utilizes a unique density-based clustering module capable of providing a full dynamical profile of clusters with varying densities. The density module (DM) is capable of detecting subtle variation in the density of mutations in 3D structures with little computation time and complexity. DM gives users the ability to detect the bridging effect, where more dense subclusters form a less dense supercluster due to a small number of mutations in between the subclusters connecting them. In addition to clustering based on physical density of mutations, DM is capable of clustering by additional biologic properties, including mutation recurrence and pathogenicity. Moreover, the underlying OPTICS algorithm utilized in the DM is inherently a “fuzzy” clustering algorithm, allowing slight variation of clusters based on the start point of the clustering algorithm from one run to another. Therefore, the DM provides a cluster membership probability measure for each mutation, giving the user the ability to choose the preferred stringency. Additionally, DM allows users to visualize the density clusters using Pymol. Since the whole dynamical set of clusters is produced by a single run, the visualization is performed so that one can reveal other clusters with higher densities by zooming in and out. Using a curated set of experimentally validated oncogenic mutations, we evaluated the performance of DM in detecting functionally activating mutations; our tool performed well in identifying driver mutations at high density thresholds. Moreover, the DM was able to identify mutations clustering at high density in BRAF, PIK3CA and KEAP1, which were not detected by other 3D clustering tools and sequence-based tools (SIFT, PolyPhen2, CHASM, etc.). The DM was also better at detecting clusters along the interface of protein-protein complexes (e.g., BRAF-KEAP1) compared to other 3D tools. In summary, HotSpot3D-DM allows for dynamical clustering, improved visualization, and identifies novel driver mutations missed by previous tools.

Citation Format: Amila Weerasinghe, Sohini Sengupta, Adam D. Scott, Maththew H. Bailey, Michael C. Wendl, Ken Chen, Gordon Mills, Li Ding. Density-based mutation clustering in 3D space [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 1306.
Abstract 132: Role of IL-17-CXCR2 axis in neutrophil recruitment facilitating breast cancer metastasis and therapy resistance
Authors: Wu Lingyun , Sharma Bhawna , Singh Rakesh K.
Published: August 2018

The major challenges for breast cancer includes therapy resistance and metastasis to distant organs. Chronic inflammation has been intimately linked with these processes. We have observed increased level of CXCR2 ligands, inflammatory immunosuppressive microenvironment, and increased metastasis in chemotherapy resistant cells. CXCR2 and its ligands have been shown to be the primary mechanism for neutrophils recruitment to the sites of primary tumors and metastases. However, their precise roles in therapy resistance and metastasis remains unclear. The aim of this project is to investigate the role of neutrophils in chemotherapy resistance and metastasis in breast cancer. In this report, we investigated the mechanisms and putative role of neutrophils in chemotherapy resistance and metastasis. We evaluated the expression level of the factors critical for neutrophil recruitments, such as interleukin (IL)-17 and its receptors (IL-17R), granulocyte colony stimulating factor (GCSF), CXCR2 and its ligands in primary tumors and metastases established from parents and chemotherapy resistant cells. The frequency of neutrophils and T-helper 17 (Th17) cells were analyzed using immunohistochemistry. Chemotherapy-resistant cell lines express higher levels of GCSF, IL-17, and IL-17R compared to Cl66 parent cells. We observed higher expression of IL-17R, CXCR2, and CXCR2 ligands in metastatic lesions compared to primary tumors. Furthermore, there were more recruitments of neutrophils and Th17 cells in resistant tumors than parent tumors. In addition, treatment of tumor cells with IL-17 significantly increased cellular proliferation together with CXCL1 and CXCL5 expression in a concentration-dependent manner. Moreover, we observed higher expression of TGFβ 2 and Th17 cell priming factors, IL-6 and IL-23, in neutrophils co-cultured with therapy resistant cells compared to the control suggesting a feed forward loop. In addition, we observed that IL17 treatment of tumor cells did not increase their tolerance to chemotherapy drug, indicating its role in establishing immune suppressive microenvironments. Together our data demonstrate an IL17-CXCR2 ligands axis induces protumorigenic inflammation, facilitating therapy resistance and metastasis through recruited neutrophils.

Citation Format: Lingyun Wu, Bhawna Sharma, Rakesh K. Singh. Role of IL-17-CXCR2 axis in neutrophil recruitment facilitating breast cancer metastasis and therapy resistance [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 132.
Abstract 1322: A systems-level approach identifies novel links between iron and fatty acid metabolism in high-grade serous ovarian cancer
Authors: Konstorum Anna , Lynch Miranda L. , Torti Suzy V. , Torti Frank M. , Laubenbacher Reinhard C.
Published: August 2018

The low survival rate for high-grade serous ovarian cancer (HGSOC) motivates novel approaches to identify causative and therapeutically targetable factors involved in HGSOC initiation and progression. Cancer cells are known to sequester iron, which can potentiate cancer progression via mechanisms that have not yet been completely elucidated. In order to uncover novel, and potentially therapeutically tractable links, between iron and HGSOC, we developed an algorithm to identify perturbed regulatory pathways containing differentially expressed iron-related genes in microarray data from clinical sources and an experimental model of HGSOC. Used in tandem with over-representation analysis for iron-related genes, the algorithm led us to uncover an iron dependence of fatty acid import and synthesis pathways, which are upregulated in HGSOC and other cancers, and to develop a network synthesizing the relationship between iron, fatty acid metabolism, and HGSOC. We use the network to derive specific hypotheses of mechanisms by which iron impacts fatty acid metabolic pathways to promote tumorigenesis. We have thus shown that a systems-level approach to identifying novel regulatory links between iron and HGSOC has yielded a previously unappreciated association between iron and fatty acid metabolism that may be exploited for therapeutic potential. This work has been supported in part by NIH grants F32CA214030 to AK, R01CA188025 to SVT, and RO1CA171101 to FMT.

Citation Format: Anna Konstorum, Miranda L. Lynch, Suzy V. Torti, Frank M. Torti, Reinhard C. Laubenbacher. A systems-level approach identifies novel links between iron and fatty acid metabolism in high-grade serous ovarian 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 1322.
Abstract 1326: Nonprotective autophagy fails to confer resistance to radiation
Authors: Xu Jingwen , Cudjoe Emmanuel Kenneth , Saleh Tareq , Patel Nipa , Alotaibi Moureq , Wu Yingliang , Lima Santiago , Gewirtz David
Published: August 2018

Radiation remains a predominant treatment option for advanced, inoperable lung cancer, however, resistance represents a major barrier against effective therapy. The exact mechanisms through which cancer cells exhibit resistance to radiotherapy still remains unresolved. Traditionally, radiation-induced autophagy is considered as a mechanism of resistance which attenuates the antitumor effects of radiotherapy i.e., autophagy plays a cytoprotective role on cell survival. To examine this concept further, we employed H460 non-small cell lung cancer (NSCLC) cells that were either p53 wild-type (p53 +/+) or p53 null (p53 -/-) to study the contribution of autophagy to radiation sensitivity. While autophagy was markedly higher in p53 +/+ cells (determined by p62 degradation, LC3B conversion, and acidic vesicle formation) the p53 +/+ cells were actually more radiosensitive than their counterpart H460 p53 -/- cells, which showed lower levels of radiation-induced autophagy. Despite modest radiosensitization by chloroquine, alternative pharmacological (3-methly adenine) and genetic (ATG5 silencing) inhibition of autophagy failed to radiosensitize either p53 +/+ or p53 -/- cells, indicating that radiation induced autophagy is not cytoprotective in function in these experimental models. Furthermore, secretome analysis (nLC-MS/MS) of p53 +/+ H460 cells showed no significant association with radioresistance-related proteins in comparison with p53 -/- H460 cells. The rate and extent of apoptosis was quite low and similar in the two cells lines, essentially ruling out apoptotic cell death as the basis for differential radiation sensitivity. Finally, Senescence was more pronounced in the p53 +/+ cells compared to the p53 -/- cells, which may contribute to the greater radiation sensitivity in the p53 +/+ cells. However, the most relevant finding in this work was that when autophagy is the nonprotective form, it does not confer inherent radiation resistance in tumor cells.

Citation Format: Jingwen Xu, Emmanuel Kenneth Cudjoe, Tareq Saleh, Nipa Patel, Moureq Alotaibi, Yingliang Wu, Santiago Lima, David Gewirtz. Nonprotective autophagy fails to confer resistance to radiation [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 1326.
Abstract 133: Synergistic effect between natural killer cells and anysomycin against hepatocellular carcinoma cells
Authors: Lee Chang Hoon , Kim Miok , Lee Kyung Won
Published: August 2018

Although advances in the clinical management of hepatocellular carcinoma (HCC) have been made, HCC is the second leading cause of cancer-related death worldwide. Currently, there are limited treatment options for advanced HCC. Therefore, novel treatment strategies for HCC are required. Here, we described a promising anti-tumor effect of anisomycin, which not only had direct killing effects but also enhanced a natural killer cell-mediated immunotherapeutic effect in HCC. To better elucidate the anti-tumor mechanisms of anisomycin, we performed a genome-scale analysis of gene expression profiles. We found that anisomycin treatment of HCC differentially regulated a broad range of immune regulation-associated genes; among them, we found that CD58, which was significantly increased on anisomycin treated HCC cells was one of the critical components for natural killer cell-mediated immunotherapeutic effect enhanced by anisomycin. Notably, this is the first report for synergistic effect of anisomycin in combination with NK cell therapy as a potential therapeutic option for HCC.

Citation Format: Chang Hoon Lee, Miok Kim, Kyung Won Lee. Synergistic effect between natural killer cells and anysomycin against hepatocellular carcinoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 133.
Abstract 1330: Role of epigenetic remodeling in sensitizing triple-negative breast cancer cells to treatment through enhanced chemotherapy-induced autophagy
Authors: Tyutyunyk Liliya , Landry Joseph , Gewirtz David , Dao Nga
Published: August 2018

Dysregulation of the epigenome is implicated in initiation and progression of variety of cancers and their acquired resistance to chemotherapy. As such targeting epigenetic regulators has the potential to modulate cancer cell biology and their sensitivity to chemotherapy and/or radiation. Our studies demonstrates that silencing of the epigenetic regulator Nucleosome Remodeling Factor (NURF) sensitizes breast tumor cells to chemotherapy and enhances the anti-tumor immune response. A screen of a variety of chemotherapeutic agents shows that NURF KD cells are selectively sensitized to Topo II inhibitors, which includes doxorubicin. NURF KD increases DNA damage (gamma H2AX staining) and autophagy (acridine orange staining) in breast tumor cells exposed to doxorubicin (Dox) and enhances growth inhibition as well as suppressing the ability of the cells to recover proliferative capacity. Increased autophagy as well as breast tumor sensitization to doxorubicin were observed using a small molecule inhibitor of NURF, suggesting that NURF can also be targeted pharmacologically. Our studies suggest that enhanced autophagy may be a primary contributor to chemo sensitivity in NURF KD cells. Studies are currently underway to confirm the role of autophagy in chemosensitization in- vitro and enhanced immune response in-vivo using autophagy deficient cells (through ATG silencing). Doxorubicin produces profound DNA damage and promotes immunogenic cell death enhanced by NURF KD. Increased cell autonomous antitumor effects by doxorubicin in concert with increased cell non-autonomous antigenicity could help to achieve tumor regression, reduce metastasis, and possibly promote long term remission in breast cancer.

Citation Format: Liliya Tyutyunyk, Joseph Landry, David Gewirtz, Nga Dao. Role of epigenetic remodeling in sensitizing triple-negative breast cancer cells to treatment through enhanced chemotherapy-induced autophagy [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 1330.
Abstract 1332: Autophagic clearance of protein aggregates is impaired in cancer cells with dysfunctional mitochondria
Authors: Biel Thomas , Rao Ashutosh
Published: August 2018

Mitophagy and aggrephagy are selective types of autophagy that degrade damaged mitochondria and protein aggregates, respectively. Dysfunctional mitochondria are known to undergo mitophagy and cause protein aggregation. How a cell can simultaneously coordinate the removal of dysfunctional mitochondria and protein aggregates via autophagy is unknown. Here, two agents, Mitoquinone (MitoQ) and MitoApocynin (MitoApo), known to induce mitochondrial dysfunction and mitophagy in MDA-MB-231 cells were used to investigate the accumulation of protein aggregates and the mechanisms that contribute to selective autophagy. Using a protein aggregate dye and a poly-ubiquitinated antibody, carbonyl cyanide m-chlorophenyl hydrazine (CCCP), MitoQ, and MitoApo were identified as agents that caused an accumulation of poly-ubiquitinated protein aggregates in cancer cells by FACS analysis and confocal imaging. Microflow imaging of the cellular lysates confirmed a relative accumulation of large protein (>460 kDa) complexes. To establish the presence of aggrephagy, the aggregates were colocalized with LC3. Wild type and ATG7 knockout MEF cells were used to test whether protein aggregates accumulated in an autophagy-dependent manner. Our data currently demonstrates the kinetics of autophagic clearance of drug-induced protein aggregation and dysfunctional mitochondria.

Citation Format: Thomas Biel, Ashutosh Rao. Autophagic clearance of protein aggregates is impaired in cancer cells with dysfunctional mitochondria [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 1332.
Abstract 1339: Development of a novel autophagy-inducing multikinase inhibitor for the treatment of castration-resistant prostate cancer
Authors: Qiao Yuanyuan
Published: August 2018

Inhibition of the androgen axis has revolutionized the treatment of metastatic castration-resistant prostate cancer (mCRPC). However, mCRPC continues to be a leading cause of cancer-related deaths in men. Here, we characterized a phase I-cleared multikinase inhibitor, ESK981, in prostate cancer preclinical models. ESK981 was markedly more potent in vitro than other kinase inhibitors, including cabozantinib and crizotinib, that have been evaluated clinically in prostate cancer. Surprisingly, we observed that ESK981 was a potent inducer of cellular vacuolization that was associated with an autophagic response. ESK981-induced autophagy could be blocked by either the depletion of the autophagy vesicle formation gene, atg5, or by autophagy inhibitors such as bafilomycin. ESK981 was further shown to induce autophagy in yeast, which is an evolutionarily conserved core cellular pathway. Moreover, ESK981 triggered prostate cancer cells to release chemokines such as CXCL10 into the cell secretome, thus suggesting that ESK981 may stimulate immune surveillance while exerting its tumor inhibitory effect. Together, ESK981 is a novel and potent autophagy-inducing multikinase inhibitor that effectively inhibits CRPC growth through the activation of the autophagic cascade.

Citation Format: Yuanyuan Qiao. Development of a novel autophagy-inducing multikinase inhibitor for the treatment of castration-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1339.
Abstract 1354: TonEBP mediates PCNA polyubiquitination on DNA damage sites via interaction with SHPRH and USP1
Authors: Kang Hyunje , Park Hyun , Yoo Eun Jin , Lee Jun Ho , Choi Soo Youn , Lee-Kwon Whaseon , Lee Kyoo-young , Hur Jin-Hoe , Seo Jeong Kon , Myung Kyungjae , Kwon Hyug Moo
Published: August 2018

PCNA ubiquitination is important for DNA damage tolerance in eukaryotic replication. While PCNA monoubiquitination activates a mutagenic translesion synthesis pathway, polyubiquitination activates an error-free template switching for DNA damage bypass. Even though PCNA polyubiquitination is critical for the maintenance of genomic integrity, underlying mechanisms are poorly understood. Here, we report that TonEBP (tonicity-responsive enhancer binding protein) is involved the PCNA polyubiquitination in response to DNA damage. TonEBP sequentially recruited an E3 ubiquitin ligase SHPRH followed by a deubiquitinase USP1 into DNA damaged sites, which showed direct correlation with dynamics of PCNA polyubiquitination. The Rel-homology domain (RHD) of TonEBP, which encircles DNA, was required for the interaction with SHRPH and USP1. Consistent with its importance for DNA damage bypass, knockdown or deletion of RHD in TonEBP increased sensitivity to methyl methanesulfonate. Collectively, TonEBP is a key player in the post-replication DNA repair.

Citation Format: Hyunje Kang, Hyun Park, Eun Jin Yoo, Jun Ho Lee, Soo Youn Choi, Whaseon Lee-Kwon, Kyoo-young Lee, Jin-Hoe Hur, Jeong Kon Seo, Kyungjae Myung, Hyug Moo Kwon. TonEBP mediates PCNA polyubiquitination on DNA damage sites via interaction with SHPRH and USP1 [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 1354.
Abstract 1356: NSMCE2 enables rescue of collapsed replication forks to prevent mitotic DNA damage
Authors: Pond Kelvin W. , DeRenty Christelle , Yagle Mary K. , Ellis Nathan
Published: August 2018

Background: Replication forks that are persistently blocked cannot restart, even after DNA damage is repaired. These forks are termed collapsed forks. In order to fully duplicate the genome, collapsed forks must be rescued by activation of nearby dormant origins of replication. Homologous recombination (HR) is essential in collapsed-fork rescue. Cells deficient in the SUMO E3 ligase NSMCE2 exhibit mitotic defects, are sensitive to DNA damaging agents, and have defects in HR. Thus, NSMCE2 regulates HR during replication stress, but the molecular mechanisms are poorly understood. In yeasts, hypomorphic NSMCE2 (MMS21) alleles accumulate HR intermediates during replication stress. We therefore hypothesized that NSMCE2 regulates RAD51 function during collapsed fork rescue.

Results: It was previously shown that sumoylation of the BLM helicase results in recruitment of RAD51 to stalled forks. Consistent with data in yeast, we found that BLM sumolyation is dependent on NSMCE2. However, contrary to our expectation, we found that the amount of RAD51 protein that accumulated at collapsed forks was over two times greater in NSMCE2-deficient cells than in normal cells. In contrast, the levels of BLM, RPA, single-stranded DNA, and γH2AX at stalled forks is reduced by half. Consistent with the low levels of γH2AX, the double-strand breaks and sister chromatid exchanges that accumulate during collapsed fork rescue were also greatly diminished in NSMCE2-deficient cells. Thus, despite the over-accumulation of RAD51 to sites of collapsed replication forks, cells are unable to perform HR efficiently, indicating that RAD51 is unable to complete its function there. In NSMCE2-deficient cells, the hyper-accumulated RAD51 at collapsed forks persists into mitosis where excess under-replicated DNA causes mitotic DNA damage.

Conclusions: The hyper-accumulation of RAD51 at stalled forks we observed in NSMCE2-deficient cells suggests that NSMCE2 is required for the remodeling of collapsed forks that normally leads to the rescue of collapsed forks, namely, the unloading of RAD51, DNA breakage, repair by HR, and completion of DNA synthesis. We suggest that the excess accumulation of RAD51 that is observed in a substantial number of cancers is not sufficient to demonstrate that the cells are HR proficient. The identification of NSMCE2 as a controller of HR-mediated fork rescue also highlights NSMCE2's potential as a new therapeutic target for combinatorial therapy of HR-dependent cancers.

Citation Format: Kelvin W. Pond, Christelle DeRenty, Mary K. Yagle, Nathan Ellis. NSMCE2 enables rescue of collapsed replication forks to prevent mitotic DNA damage [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 1356.
Abstract 1358: Translational regulation of DNA repair: The mechanism under cap-independent translational regulation of RPA2
Authors: Cui Jia-Jia , Wang Lei-Yun , Guo Ao-Xiang , Yin Ji-Ye
Published: August 2018

Purpose: Maintenance of genome integrity is critical for any living cells. A variety of endogenous and exogenous factors can cause DNA damage. RPA2 (Replication protein A2) is a kind of single-stranded DNA binding protein complexes and play an important role in the DNA repair. Internal ribosome entry site (IRES) element is a RNA sequence with a complex structure, the activation of RPA2 IRES element can cause its abnormal expression and finally effects DNA repair pathway. IRES is regulated by eukaryotic translation initiation factors (eIFs) and IRES trans-acting factors (ITAFs). UNR (upstream of N-ras), as one of the important IRES trans-acting factors, and eIF3a (Eukaryotic initiation factor 3a) as one of the important eIFs play important roles in the regulation of RPA2 protein expression via IRES elements. We want to explore the role and potential mechanism of translational regulation of UNR and eIF3a in DNA repair. Methods: In the current study, biotin pull down assay was taken to investigate the interaction between RPA2 IRES and UNR. Western blot and qPCR were used to detect protein and mRNA level respectively. CO-IP assay were conducted for interaction of eIF3a with UNR. Immunofluorescence assay was taken to investigate the intracellular localization of UNR and eIF3a, as well as the regulation of UNR on DNA double strand break repair pathway. GST pull down assay was carried out to further identify the interaction between two proteins. Electrophoretic mobility shift assay was carried out to explore the domains of UNR and eIF3a that bind to RPA2 IRES element. Dual-luciferase reporter assay, comet assay and immunofluorescence assay were used to investigate the DNA repair pathway activity. Results: UNR and eIF3a translationally regulate RPA2 by combining with RPA2 IRES. It was found that the expression of RPA2 was down-regulated by UNR with no change on mRNA level. We further found that UNR combined with RPA2 IRES via CSD1 domain. And no domains of eIF3a bind to RPA2 IRES RNA.Meanwhile, the location of UNR and eIF3a was consistent. A series of assays indicate that CSD1 domain of UNR bind to PCI domain of. RPA2 play an important role in the DNA repair. We found that UNR down-regulated double-strand break repair (DSB) activity and nucleotide excision repair (NER) activity. Conclusion: UNR and eIF3a can regulate the expression of RPA2 on translational level, and further regulate the activity of DNA repair, which may reveal the potential mechanism of translational regulation and DNA repair.

Citation Format: Jia-Jia Cui, Lei-Yun Wang, Ao-Xiang Guo, Ji-Ye Yin. Translational regulation of DNA repair: The mechanism under cap-independent translational regulation of RPA2 [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 1358.
Abstract 1360: A PARylation biosensor genetic screen to identify novel PARylation targets
Authors: Krastev Dragomir B. , Pettitt Stephen J. , Campbell James , Ashworth Alan , Lord Christopher J.
Published: August 2018

Poly(ADP-ribosylation) (PARylation) is a reversible post-translational protein modification that regulates a number of cellular processes. It is generated by a family of PAR polymerases (PARP), of which PARP1 has recently been exploited as a target in ovarian cancer carrying BRCA1/2 mutations. In addition, tankyrase (PARP5A and B), with its regulation of Wnt signalling, telomeres and the mitotic apparatus, has attracted much effort in the development of targeted therapies. All this necessitates the identification of PARylation targets and their mode of regulation by the modification. Genetically encoded fluorescent biosensor that can report the PARylation state of a target protein has the potential to report the modulation of the PARylation target in living cells. We used PAR-binding PBZ domains fused to a non-fluorescent fragment of a GFP molecule in combination with a protein of interest fused to the complementing fragment of GFP. When this protein is PARylated, it is recognized by the PBZ domain, which brings in proximity the two GFP fragments, and restores the GFP fluorescence. Known targets of PARylation, e.g. TERF1 and AXIN1, showed specific spatial patterns of PARylation validating our approach. In order to identify novel PARylation targets, we have designed a genetic screen based on the biosensor. We introduced the GFP fragment in a transposon, which was randomly integrated in cellular genome, thus generating cellular libraries of thousands of tagged genes. The introduction of the PAR binding probe lead to GFP fluorescence in clones that harbour putative PARylation targets. We developed a next generation sequencing approach to isolate the transposon integration sites and annotated the target genes. Reassuringly, we have identified NPM1, a well-known PARylation target, as a strongly scoring gene. We focused on one of novel identified hits (CTIF). We showed that CTIF is PARylated in vivo in a tankyrase-dependent manner. Furthermore, the biosensor revealed that this PARylation is predominant in a specific cellular compartment surrounding the centrosome. Functional studies have implicated this protein the regulation of the centrosomal satellites. Collectively, we are presenting a new approach to identify PARylated proteins and study their patterns of PARylation and regulation in vivo.

Citation Format: Dragomir B. Krastev, Stephen J. Pettitt, James Campbell, Alan Ashworth, Christopher J. Lord. A PARylation biosensor genetic screen to identify novel PARylation targets [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 1360.
Abstract 1365: NR1D1 enhances reactive oxygen species-induced DNA damage by inhibition of PARP1
Authors: Ka Na-Lee , Na Tae-Young , Lee Mi-Ock
Published: August 2018

Breast cancer is the most commonly diagnosed cancer in women worldwide. Cancer cells produce higher levels of intracellular reactive oxygen species (ROS) than their normal counterparts, because of their accelerated metabolism, mitochondrial dysfunction, and antioxidant deficit. The oxidative stress in cancer cells may provide clinical benefits, which can be associated with a better response to anticancer therapies. Therefore, identifying the regulatory pathway of oxidative stress in cancer cells is important in the development of therapeutic targets that enhance sensitivity to ROS-generating anticancer therapies. In this study, we identified that nuclear receptor subfamily 1, group D, member 1 (NR1D1; Rev-erbα) inhibited DNA repair of ROS-induced DNA damage in breast cancer cells. NR1D1 interacted with poly(ADP-ribose) polymerase 1 (PARP1) and subsequently inhibited catalytic activity of PARP1. NR1D1 enhanced accumulation of oxidative DNA damage, which increased sensitivity of breast cancer cells to the ROS-induced cell death. Our findings suggest that NR1D1 may provide better therapeutic options for breast cancer treatment, especially in those patients treated with ROS-inducing chemotherapeutic agents.

Citation Format: Na-Lee Ka, Tae-Young Na, Mi-Ock Lee. NR1D1 enhances reactive oxygen species-induced DNA damage by inhibition of PARP1 [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 1365.
Abstract 138: PRMT6 promotes lung tumor growth via the modulation of macrophage M2 polarization
Authors: Avasarala Sreedevi , Wu Pei-Ying , Khan Samia Q. , Scoyk Michelle Van , Su Yanlin , David Odile , Gupta Vineet , Bedford Mark T. , Winn Robert A. , Bikkavilli Rama Kamesh
Published: August 2018

Lung cancer kills more people in the United States than any other malignancy. High mortalities are in part due to the diagnosis at an advanced inoperable stage where the 5-year survival is only 4%. Therefore, improving survival will require increased knowledge of genes that drive the complex mechanisms of lung cancer initiation and progression. Emerging data implicate novel roles for protein arginine methylation in lung cancers. In protein arginine methylation, nitrogen/s of arginine can be post-translationally modified via the addition of a methyl group, catalyzed by a class of enzymes, protein arginine methyl transferases (PRMTs). Based on the computational analysis, tissue staining, and immunoblotting we have identified an increased expression of PRMT6 in lung tumors. To test the biologic relevance of PRMT6 in lung tumor development, we employed an inducible lung-targeted PRMT6 gain-of-function (GOF) mouse model. For the first time, we show that lung-targeted overexpression of PRMT6 results in spontaneous lung tumor development. We also observed a robust increase in lung tumors in the PRMT6 GOF mice in response to urethane, a KRas mutagen, and a prototypical model to study lung tumorigenesis. Excitingly, we detected an increase in the number of tumor-associated macrophages (TAMs) in the PRMT6 GOF mice. Further characterization of the TAMs from the PRMT6 GOF mice revealed that they were M2 polarized with immunosuppressive, angiogenesis-enhancing, and tumor-promoting activities. Taken together, these results demonstrate a novel role for PRMT6 in lung tumor development by way of macrophage M2 polarization. Therefore, defining the complete mechanism of PRMT6-mediated macrophage M2 polarization could lead to the development of novel therapeutic strategies to treat lung cancer.

Citation Format: Sreedevi Avasarala, Pei-Ying Wu, Samia Q. Khan, Michelle Van Scoyk, Yanlin Su, Odile David, Vineet Gupta, Mark T. Bedford, Robert A. Winn, Rama Kamesh Bikkavilli. PRMT6 promotes lung tumor growth via the modulation of macrophage M2 polarization [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 138.
Abstract 1380: Towards peptidomimetics to target DOT1L recruitment in MLL-AF9 leukemia
Authors: Grigsby Sierrah Marie , Chase Jennifer , Waas Bridget , Friedman Ann , Du Lei , Yao Aihong , Ropa James , Serio Justin , Muntean Andrew , Maillard Ivan , Sun Haying , Nikolovska-Coleska Zaneta
Published: August 2018

Leukemias harboring rearrangements of mixed-lineage leukemia gene (MLL1) are associated with poor clinical outcomes, and new therapeutic approaches are needed. Rearrangement of the MLL1 gene generates fusion oncoproteins which drive the high expression of the clustered homeobox (HOX) genes and induce leukemic transformation. Genome-wide histone methylation studies have revealed that the abnormal expression of MLL1 fusion target genes is associated with high levels of histone H3 lysine 79 (H3K79) methylation. Recruitment of DOT1L (disruptor of telomeric silencing 1-like), a unique histone methyltransferase that catalyzes methylation of H3K79, proved to be essential for the transforming activity of multiple MLL fusion proteins. We have mapped the binding site to a short segment of 10 amino acids in DOT1L and shown that DOT1L mutants lacking these residues did not support transformation by MLL-AF9. We hypothesized that by targeting the AF9-DOT1L protein-protein interactions (PPIs), we would selectively kill MLL-AF9 cells without effecting DOT1L role in normal hematopoiesis. Using established DOT1Lf/f MLL-AF9 with reintroduced WT-DOT1L, DOT1L missing 10aa AF9-binding domain (D10), DOT1L with a point mutation in the AF9-binding domain (I867A) and enzymatically inactive DOT1L (RCR), we were able to demonstrate that by disrupting the AF9-DOT1L PPIs, although we can inhibit leukemogenesis similarly to enzymatic inhibition, this interaction is not essential for normal hematopoiesis. Based on our initial studies to map the DOT1L interaction site and in conjunction with utilizing reported NMR structures of the AF9-DOT1L complex, we investigated the nature of the interactions and the minimum length of the peptide. Using different natural and unnatural amino acids, we successfully designed a 7mer peptide with KD of 10 nM and 25 nM against AF9 and ENL, respectively, showing similar potency as the originally identified and validated 10mer peptide. These results lay the groundwork for further optimization of the 7mer peptide towards developing DOT1L peptidomimetics with improved potency and cellular activity, to further validate the PPIs between DOT1L and MLL-fusion proteins as a potential therapeutic target for MLL rearranged leukemia.

Citation Format: Sierrah Marie Grigsby, Jennifer Chase, Bridget Waas, Ann Friedman, Lei Du, Aihong Yao, James Ropa, Justin Serio, Andrew Muntean, Ivan Maillard, Haying Sun, Zaneta Nikolovska-Coleska. Towards peptidomimetics to target DOT1L recruitment in MLL-AF9 leukemia [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 1380.