bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3445: Mechanism, physiological and therapeutic implications of LGR-independent potentiation of WNT signaling by R-spondins
Authors: Lebensohn Andres M. , Rohatgi Rajat
Published: August 2019

The WNT signaling pathway encodes positional information in animals, orchestrates patterning and morphogenesis during embryonic development, and promotes tissue renewal and regeneration in adults. Dysregulation of WNT signaling has been implicated in many diseases, including developmental malformations, skeletal and dental abnormalities, cardiovascular and neurodegenerative disorders, diabetes, and many types of cancer. Members of the R-spondin family of secreted growth factors have emerged as key regulators of WNT signaling strength. The R-spondin system is a vertebrate adaptation that may be related to the appearance of sophisticated stem cell compartments, which require exquisite control of WNT signaling. The four members of the family (R-spondin 1-4) can strongly potentiate responses to WNT ligands during development and in stem cells, but the mechanisms by which they transduce signals are not fully understood and the reasons why they produce different physiological effects are largely unknown. Unexpectedly, we discovered that R-spondins 2 and 3 can uniquely potentiate WNT signaling in cells lacking LGRs 4, 5 and 6, the principal R-spondin receptors. We determined the protein domains on R-spondins necessary and sufficient for this new mode of signaling, and showed that it is mediated by an alternative interaction with cell-surface heparan sulfate proteoglycans. This finding is transformative because LGRs 4-6 were thought to be required to transduce all R-spondin signals and hence determine their site of action. Indeed, these LGRs are highly expressed in many stem cell compartments whose maintenance depends on potentiation of WNT signaling by R-spondins. Instead our work shows that there are two modes of signaling by R-spondins, an LGR-dependent and an LGR-independent mode, and that different R-spondins may use distinct molecular mechanisms to transduce signals in various biological contexts. Supporting our findings, recent work from another group demonstrated that during limb development, R-spondin 2 signals through an LGR-independent mechanism. Since components of the R-spondin signaling module are commonly mutated in cancer, therapeutic targeting of LGR-independent signaling deserves evaluation. I will present the results demonstrating our conclusions and discuss their implications in physiology and disease.

Citation Format: Andres M. Lebensohn, Rajat Rohatgi. Mechanism, physiological and therapeutic implications of LGR-independent potentiation of WNT signaling by R-spondins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3445.
Abstract 3448: Dasatinib attenuates Src pathway signaling and combined with veliparib and carboplatin potently inhibits tumor growth in a preclinical model of TNBC
Authors: Sun Yuliang , Lin Xiaoqian , Aske Jennifer C. , Williams Casey , Abramovitz Mark , Leyland-Jones Brian
Published: August 2019

Background: The combination of a poly ADP ribose polymerase (PARP) inhibitor with a DNA-damaging agent has shown promise in treating triple-negative breast cancer (TNBC); however, not all patients respond to this combination (Rugo HS et al. The New England Journal of Medicine. 2016). The Src protein kinase modulates multiple cancer cell properties and plays a key role in tumorigenic processes. However, Src inhibitors as single agents have shown limited effects in solid tumors (Finn RS et al. Clinical Cancer Research. 2011). In this study, we examined the antitumor effects of the Src inhibitor dasatinib, the PARP inhibitor veliparib, and the DNA-damaging agent carboplatin in TNBC cells in vitro and tumors in vivo to try to identify the combination with the most clinical potential.

Methods: The KRAS/BRAF-mutated TNBC cell line, MDA-MB-231, was used in this study. For in vitro studies, proliferation, cellular apoptosis, and 3D on-top clonogenic growth were measured, tube formation assays and western blot analysis were performed. For in vivo studies, a xenograft model was used to examine treatment responses, and immunohistochemical analysis was performed.

Results: Surprisingly, treatment with the combination of veliparib plus carboplatin led to an increase in Src phosphorylation. Importantly, addition of dasatinib attenuated Src overexpression induced by veliparib plus carboplatin and further inhibited the downstream signaling of Src, thereby enhancing the antitumor efficacy of veliparib plus carboplatin. In an MDA-MB-231 xenograft model, the triple combination of dasatinib with veliparib plus carboplatin showed greater tumor growth inhibitory effects compared with single agents or double combinations. No systemic toxicity was observed in mice treated with the triple combination.

Conclusions: Here, we provide novel evidence that veliparib combined with carboplatin drives Src activation in the preclinical model tested. Moreover, we provide provocative in vitro and in vivo data highlighting the potential for increasing therapeutic efficacy by combining dasatinib with veliparib and carboplatin in patients with KRAS/BRAF-mutated TNBC.

Citation Format: Yuliang Sun, Xiaoqian Lin, Jennifer C. Aske, Casey Williams, Mark Abramovitz, Brian Leyland-Jones. Dasatinib attenuates Src pathway signaling and combined with veliparib and carboplatin potently inhibits tumor growth in a preclinical model of TNBC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3448.
Abstract 3471: Mutual regulation of MDM4 and TOP2A in cancer cell proliferation
Authors: Liu Tao , Zhang Hailong , Yi Sha , Gu Lubing , Zhou Muxiang
Published: August 2019

MDM4 and topoisomerase IIα (TOP2A) are important cancer-related factors. MDM4 acts as an oncoprotein promoting cancer progression by inhibiting tumor suppressor p53. As a DNA replication- and cell division-regulating enzyme, TOP2A is the main target of many anticancer therapy regimens, even though the exact role of TOP2A in cancer remains elusive. Herein we report that MDM4 and TOP2A bind to each other and are mutually upregulated at the post-translational level, leading to enhanced TOP2A catalytic activity, inhibition of p53, and increased tumor-cell proliferation. We demonstrate that the C-terminal region (CTR) of TOP2A binds to a unique sequence (residues 188-238) of MDM4, which contains an auto-inhibitory segment regulating MDM4-p53 interaction. TOP2A binding in turn activates MDM4 for p53 binding, resulting in enhanced inhibition of p53 and cancer cell proliferation. Conversely, binding of the MDM4 sequence to the CTR of TOP2A upregulates TOP2A protein expression. This in turn interferes with TOP2A DNA-decatenating activity. These results reveal novel functions of MDM4 and TOP2A as well as their interactions in oncogenesis, suggesting that inhibition of the MDM4-TOP2A interaction may represent a novel strategy in specifically and simultaneously targeting TOP2A and MDM4 for cancer treatment.

Citation Format: Tao Liu, Hailong Zhang, Sha Yi, Lubing Gu, Muxiang Zhou. Mutual regulation of MDM4 and TOP2A in cancer cell proliferation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3471.
Abstract 3486: Combined inhibition of tumor suppressors PTEN and PP2A drives anoikis resistance and is associated with therapy relapse in prostate cancer
Authors: Rupp Christian , Isomursu Aleksi , Aakula Anna , Erickson Andrew , Li Song-Ping , Kaur Amanpreet , Shah Pragya , Pokharel Yuba R. , Trottman Lloyd , Lammerding Jan , Rannikko Antti , Taimen Pekka , Mirtti Tuomas , Paatero Ilkka , Ivaska Johanna , Westermarck Jukka K.
Published: August 2019

Reactivation of tumor suppressor phosphatases may provide entirely novel opportunities for cancer therapy. Here, we discover clinically relevant functional co-operation between loss of activities of two human tumor suppressor phosphatases, PTEN, and PP2A. Analysis of prostate cancer tissue microarray material consisting of 358 patients treated primarily with radical prostatectomy revealed that overexpression of PP2A inhibitor protein PME-1 associates with significantly shorter time to therapy relapse in patients with PTEN-deficient PrCa. Further, PP2A inhibition by PME-1 overexpression in PTEN-deficient cell models inhibits apoptosis induction in anchorage-independent conditions (anoikis). PP2A reactivation by small molecules (SMAPs) was also found to inhibit viability of PTEN-deficient PrCa cells. Importantly, rather than regulating the well-known PP2A target pathways, PME-1 was found to physically associate with, and to regulate deformability of the nuclear lamina in PrCa cells. Mass spectrometry phosphoproteomics analysis identified several PME-1-regulated nuclear lamina constituents, and PME-1 deficient cells with compromised nuclear lamina were particularly vulnerable to apoptosis induction by mechanical stress. As a direct molecular target, Lamin A/C phosphorylation was found to be protected by PME-1-mediated PP2A inhibition under anoikis-inducing conditions. PME-1 inhibition in PrCa cells resulted in increased apoptosis in an in ovo tumor model, and PME-1-depleted cells had compromised long-term survival in zebrafish circulation. In summary we discover that PP2A reactivation by PME-1 targeting sensitizes PTEN-deficient PrCa cells to anoikis. Clinically, the results identify PME-1 as a novel candidate biomarker for increased relapse risk in PTEN-deficient PrCa, and indicate pharmacological PP2A activation as a novel potential therapeutic approach against circulating prostate cancer cells. At the general level, the results clearly emphasize the need for better understanding of phosphatases as key modulators of cancer progression.

Citation Format: Christian Rupp, Aleksi Isomursu, Anna Aakula, Andrew Erickson, Song-Ping Li, Amanpreet Kaur, Pragya Shah, Yuba R. Pokharel, Lloyd Trottman, Jan Lammerding, Antti Rannikko, Pekka Taimen, Tuomas Mirtti, Ilkka Paatero, Johanna Ivaska, Jukka K. Westermarck. Combined inhibition of tumor suppressors PTEN and PP2A drives anoikis resistance and is associated with therapy relapse in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3486.
Abstract 3499: Dianhydrogalactitol (VAL-083) in combination with AZD1775 increases survival in diffuse intrinsic pontine glioma (DIPG), in vivo
Authors: Steino Anne , Yang Xiaodong , Kline Cassie , Bacha Jeffrey , Brown Dennis M. , Mueller Sabine
Published: August 2019

OBJECTION: VAL-083 is a structurally unique bi-functional DNA targeting agent that readily crosses the blood-brain barrier and accumulates in brain tumor tissue. VAL-083 induces interstrand crosslinks leading to DNA double-strand breaks and S/G2 cell cycle arrest. By inhibiting the G2 checkpoint regulator kinase Wee1, AZD1775 allows a cancer cell with DNA damage to progress past the G2 checkpoint and into premature mitosis leading to cancer cell death. Herein we assess the activity of VAL-083 as single agent as well as in combination with Wee1 Kinase inhibitor AZD1775 in patient derived model systems of DIPG.

METHODS: DIPG derived cell lines SF8628 and NEM157 (H3.3K27) as well as SF10693 (H3.1K27M) and pediatric glioblastoma cell lines SF188 (H3.3K27 wildtype) were treated with increasing concentrations of single agent VAL-083 as well as in combination with AZD1775. To determine potential synergistic activity, we applied the Chou-Talalay method, which allows the quantitative determination of drug interactions by calculating a combination index (CI). In vivo activity of VAL-083 (3 mg/kg) as single agent as well as in combination with AZD1775 (60 mg/kg) was assessed in an orthotopic engraftment model of pediatric DIPG (SF8628).

RESULTS: The IC50 of VAL-083 as single agent ranged from 1µM to 10µM. VAL-083 exhibited synergistic activity in combination with AZD1775 in cell lines SF8628 and SF188 and additive effect in NEM157 with CI values ranging from 0.04 to 0.95, with CI < 1 indicating synergy. In vivo, treatment with VAL-083 as single agent and in combination with AZD1775 conferred significant survival benefit to mice with engrafted DIPG tumors compared to control as well as single agent treatment with AZD1775. The median survival for mice treated with VAL-083 alone was 54.5 days (p=0.0004, VAL-083 vs. control) and for the VAL-083/AZD1775 combination it was 62 days (p<0.0001, VAL-083/AZD1775 vs. control), compared to 44 days for control and 47 days for mice treated with AZD1775 alone (p=0.0839, AZD1775 vs. control).

CONCLUSION: Our present study highlights that the combination of VAL-083 and AZD1775 might be a promising new therapeutic strategy for children with DIPG. Ongoing studies continue to assess the in vivo activity in other DIPG models as well explore the underlying mechanism of action of the combination strategy.

Citation Format: Anne Steino, Xiaodong Yang, Cassie Kline, Jeffrey Bacha, Dennis M. Brown, Sabine Mueller. Dianhydrogalactitol (VAL-083) in combination with AZD1775 increases survival in diffuse intrinsic pontine glioma (DIPG), in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3499.
Abstract 3525: Achieving reproducibility and accuracy in cancer mutation detection with whole-genome and whole-exome sequencing
Authors: Xiao Wenming ,   The Somatic Mutation Working Group of the SEQC-IIConsortium
Published: August 2019

Clinical applications of precision oncology require accurate tests that can distinguish tumor-specific mutations from errors introduced at each step of next generation sequencing (NGS). For NGS to successfully improve patient lives, discriminating between true mutations and artifacts is crucial. We systematically interrogated somatic mutations in paired tumor normal cell lines to identify factors affecting detection reproducibility and accuracy. Different types of samples with varying input amount and tumor purity were processed using multiple library construction protocols. Whole-genome and whole-exome sequencing were carried out at six sequencing centers followed by processing with nine bioinformatics pipelines to evaluate their reproducibility. We identified artifacts due to sample and library processing and evaluated the capabilities and limitations of bioinformatics tools for artifact detection and removal. By examining the interaction and effect of various wet lab and computational parameters concomitantly, here we recommend actionable best practices for mutation detection in clinical applications using NGS technologies.

Citation Format: Wenming Xiao, The Somatic Mutation Working Group of the SEQC-IIConsortium. Achieving reproducibility and accuracy in cancer mutation detection with whole-genome and whole-exome sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3525.
Abstract 3526: Incorporation of unique molecular identifiers (UMIs) into unique dual sample indexing (UDI) improves the accuracy of quantitative next generation sequencing
Authors: Liu Pingfang , Krishnan Keerthana , Devoe Camille X. , Langhorst Bradley W. , Dimalanta Eileen , Davis Theodore B.
Published: August 2019

Accurate analysis of quantitative NGS data is critical for low frequency variant detection, identification of differentially expressed transcripts, and correct diagnosis and patient care in a clinical NGS setting. Two major factors affecting sequencing accuracy are 1) PCR duplication arising from amplification of library molecules; and 2) errors introduced during library preparation and actual sequencing on the flow cell. Standard practice for identification and removal of PCR duplicates relies on aligning reads to the same genomic coordinates. However, this approach can not differentiate between PCR duplicates and reads originating from unique molecules with identical ends. The most effective method for error correction is Duplex Sequencing, which utilizes UMIs to tag both strands of each individual DNA duplex followed by building consensus sequences. Unfortunately, this approach is labor intensive and cost-prohibitive for complex genomes and large target panels. Therefore, a simple and reliable approach for duplicate removal and error correction would greatly facilitate the wider adoption of NGS technology for diagnostics and clinical applications.

In this study, we incorporate UMIs into UDI systems and assess the effect on the accuracy of quantitative sequencing assays. We first study the effectiveness of various computational methods to account for UMIs and remove base-calling errors introduced during sequencing. We then analyze the utility of UMIs for 1) unique and low abundance transcript identification and accurate transcript quantification; and 2) error correction in low frequency variant detection in genomic sequencing from both high quality cell line DNA and low quality FFPE DNA. In addition, we demonstrate that combining unique dual sample indexing with UMI molecular barcoding further improves data analysis accuracy, especially on patterned flow cells. Our approach involves a simple new UMI-containing UDI adaptor design that can also be applied to other sequencing methods and platforms.

Citation Format: Pingfang Liu, Keerthana Krishnan, Camille X. Devoe, Bradley W. Langhorst, Eileen Dimalanta, Theodore B. Davis. Incorporation of unique molecular identifiers (UMIs) into unique dual sample indexing (UDI) improves the accuracy of quantitative next generation sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3526.
Abstract 3531: Modular RNA expression analysis using whole transcriptome and targeted analysis in single cells on BD Rhapsody
Authors: Nakamoto Margaret , Lam Gretchen , Chang Christina , Shum Eleen
Published: August 2019

Advances in genomics have allowed high-throughput single-cell whole transcriptome analysis (WTA) to enable the discovery of biomarkers and cellular pathways that may be important in diseases. As a complementary tool to WTA, targeted analysis can provide high sensitivity of single cell gene expression data with lower sequencing cost. Here we demonstrate using the BD Rhapsody platform in a modular fashion to generate both WTA and targeted sequencing libraries from the same sample. Utilizing the archivability of the BD Rhapsody beads, we first subsampled a portion of the beads to generate WTA data, from which we identified genes that are important for T cell exhaustion. We then designed a targeted panel to profile the rest of the beads from the same experiment. The study demonstrates the versatility of the BD Rhapsody single cell platform and the ability to conduct different workflows on the same sample. This approach offers flexibility and choice for users in experimental design and allows users to easily transition from novel target discovery to robust data validation using targeted panels.

Citation Format: Margaret Nakamoto, Gretchen Lam, Christina Chang, Eleen Shum. Modular RNA expression analysis using whole transcriptome and targeted analysis in single cells on BD Rhapsody [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3531.
Abstract 3533: Low-frequency variant detection in cell-free DNA by integrating double-stranded unique molecular identifiers with ultrafast amplicon-based library preparations
Authors: Lee Lucie S. , Liu Yang Lily , Pendleton Kathryn , Liu Jeffrey , Lin Lifeng Phil , Liu Guoying , Liu Zhitong
Published: August 2019

The use of liquid biopsy has seen tremendous growth in recent years as cell-free DNA (cfDNA) is a noninvasive and easily obtainable sample type with many diagnostics and prognostic values. While liquid biopsy can potentially enable new applications, such as early cancer detection, treatment monitoring, and drug resistance screening, it also presents challenges to accurate variant detection due to the low fraction of mutant DNA present in theses samples, which can be easily buried by the artifacts and background noise from systematic PCR and sequencing errors. While some success has been made using adapters with unique molecular identifiers (UMIs) in hybrid capture-based methods and performing deep sequencing, such approaches suffer from long, tricky, and tedious workflows and disappointingly low on-target rates. Additionally, the large amount of cfDNA required for hybrid capture workflows is not realistic to obtain from patients.

We have developed the CleanPlex® UMI technology to provide a fast, simple, and reliable NGS solution for low-frequency variant detection. The technology features three simple steps to generate molecular-barcoded and target-enriched NGS libraries in under 4 hours. This amplicon-based method consists of a multiplex PCR reaction for molecular barcoding, followed by a biochemical reaction for removal of redundant PCR products, and finally a second round of PCR to add Illumina adapter sequences and sample indexes. To validate this new method, we designed an NGS panel targeting frequently mutated hotspots in 23 genes associated with lung cancer. NGS libraries were prepared with commercial reference cfDNA at 0.1% to 0.5% minor allele frequencies (mAF) and sequenced on an Illumina NextSeq®.

Overall, we obtain high detection sensitivity for low-frequency alleles even at low DNA inputs. When including UMI in the analysis, we observe significant reductions in false positive calls. Nearly all known mutations covered by the CleanPlex UMI panel (12 out of 13) are detected at 0.1% mAF using 50ng of the reference cfDNA. At 0.25% mAF, all known mutants are detected with 100% PPV using 50ng of DNA. With only 20 ng of cfDNA, the method can achieve the same 100% detection at 0.5% mAF. The CleanPlex® UMI technology demonstrates high sensitivity with low false positive rate, for the detection of low-frequency alleles.

Citation Format: Lucie S. Lee, Yang Lily Liu, Kathryn Pendleton, Jeffrey Liu, Lifeng Phil Lin, Guoying Liu, Zhitong Liu. Low-frequency variant detection in cell-free DNA by integrating double-stranded unique molecular identifiers with ultrafast amplicon-based library preparations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3533.
Abstract 3547: MicroRNA-496 inhibits triple negative breast cancer cell proliferation by targeting DEL-1
Authors: Baek Dong Won , Jeong Jae-Hwan , Lee Soo Jung , Lee Jiyeon , Chae Yee Soo , Kim Wan WooK , Kang Jieun , Park Ho Yong , Jung Jin Hyang , Jeong Ji Yun , Park Ji Young , Park Keon Uk
Published: August 2019

Background and Objectives: Del-1 is linked to the pathogenesis of various cancers including breast cancer; however, the regulation of Del-1 expression remains unclear. The current study investigated how microRNA-496 (miR-496) regulates Del-1 expression in triple negative breast cancer (TNBC).

Methods: Del-1 mRNA and miR-496 were measured by quantitative PCR in breast cancer cells (MDA-MB-231, MCF7, SK-BR3, and T-47D) and tissues from 30 patients with TNBC. The effects of miR-496 on cell proliferation, migration, and invasion were determined in MTT, wound healing, and Matrigel Transwell assays, respectively.

Results: In MDA-MB-231, miR-496 levels were remarkably low and Del-1 mRNA was higher compared to other breast cancer cell lines. Luciferase reporter assays revealed that miR-496 binds the 3′-UTR of Del-1 and that Del-1 expression is downregulated by miR-496 mimics. Furthermore, miR-496 inhibited the proliferation, migration, and invasion of MDA-MB-231 cells. The effects of miR-496 on cell proliferation were additive with those of miR-137, another miRNA that regulates Del-1 expression. Moreover, in the 30 TNBC specimens, miR-496 was downregulated (P < 0.005) and the levels of Del-1 in the plasma was significantly elevated as compared to normal controls (P = 0.0142). TCGA data showed the correlation of miR-496 expression with better overall survival in patients with early TNBC.

Conclusions: In in silico and in vitro analyses, we showed that Del-1 is a target of miR-496 in TNBC and thereby affects cancer progression. Our findings suggest that miR-496 and Del-1 might act as modulating factors in TNBC and are new biomarkers for patients with TNBC.

Citation Format: Dong Won Baek, Jae-Hwan Jeong, Soo Jung Lee, Jiyeon Lee, Yee Soo Chae, Wan WooK Kim, Jieun Kang, Ho Yong Park, Jin Hyang Jung, Ji Yun Jeong, Ji Young Park, Keon Uk Park. MicroRNA-496 inhibits triple negative breast cancer cell proliferation by targeting DEL-1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3547.
Abstract 3550: microRNA-211 promotes aggressive melanoma growth in vivo by epigenetic modification, and contributes to BRAFV600E inhibitor resistance via ERK5 signaling
Authors: Lee Bongyong , Sahoo Anupama , Sawada Junko , Zisoulis Dimitrios G. , Marchica John , Sahoo Sanjay , Layng Fabiana I Alves De Lima , Finlay Darren , Mazar Joseph , Joshi Piyush , Komatsu Masanobu , Vuori Kristiina , Powis Garth , Jong Petrus R. de , Ray Animesh , Perera Ranjan J.
Published: August 2019

The microRNA miR-211 is an established participant in melanomagenesis, but controversy exists as to whether it acts as a bone fide tumor suppressor or oncogene. Here we ectopically expressed miR-211 in the BRAF v600E-mutant A375 melanoma cell line and examined its effect in xenografts in vivo. The miR-211 ectopic expression promoted aggressive tumor xenograft growth with extensive cell proliferation, and angiogenesis. ChIP-seq and single cell sequencing analysis of xenograft tissues demonstrated that aggressive tumor formation is partly associated with H3K27me3 and H3K4me3, and migration of cells from mouse tissues to tumor locus. Interrogation of xenograft transcriptomics data revealed activation of the ERK5 pathway, itself negatively regulated by miR-211 target genes, BIRC2 and DUSP6, further confirmed as direct miR-211 target genes by RNA immunopurification with RNA-seq (RIP-seq) and site-directed mutagenesis. miR-211 conferred resistance to the BRAF inhibitor vemurafenib, and MEK inhibitor cobimetinib with corresponding increases in ERK5 phosphorylation. The miR-211-ERK5 axis may represent a novel therapeutic target, but however, miR-211 is exquisitely pleiotropic in the complex in vivo tumor environment and its context must be considered carefully in diagnostic and therapeutic development.

Citation Format: Bongyong Lee, Anupama Sahoo, Junko Sawada, Dimitrios G. Zisoulis, John Marchica, Sanjay Sahoo, Fabiana I Alves De Lima Layng, Darren Finlay, Joseph Mazar, Piyush Joshi, Masanobu Komatsu, Kristiina Vuori, Garth Powis, Petrus R. de Jong, Animesh Ray, Ranjan J. Perera. microRNA-211 promotes aggressive melanoma growth in vivo by epigenetic modification, and contributes to BRAFV600E inhibitor resistance via ERK5 signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3550.
Abstract 3557: miR-181a is a key driver of genomic instability and ovarian cancer tumorigenesis through the regulation of RB1
Authors: Knarr Matthew , Kwiatkowski Lily , Nagaraj Anil , Drapkin Ronny , DiFeo Analisa
Published: August 2019

Epithelial ovarian cancer (EOC) is one of the deadliest gynecological cancers currently diagnosed in women with approximately 20,000 new cases and 15,000 deaths per year. Ovarian cancer can be stratified into distinct subtypes of which high-grade serous ovarian cancer (HGSOC) comprises 90% of all cases and has the highest mortality rate. Reducing the high mortality rate associated with HGSOC currently has two primary barriers: 1) most HGSOCs are diagnosed at a post-metastatic stage and 2) up to 70% of patients recur within 5 years. Both of these limitations are due to the fact that the drivers of tumor initiation are poorly understood. In order to change the status quo of the last 30 years, a new approach that focuses on understanding the origins of HGSOC is required. Our lab has recently discovered that miR-181a promotes key hallmarks of HGSOC progression in fallopian tube secretory epithelial cells (FTSECs) which are precursor cells of HGSOC. We show that enhanced expression of miR-181a in FTSECs results in increased proliferation, survival, and anchorage independent growth in vitro. Furthermore, FTSECs with miR-181a overexpression are able to form tumors in vivo whereas FTSECs that overexpress a control scramble miRNA cannot. Using cell cycle analysis, dynamic live-cell imaging, and SNP array analysis, we also found that miR-181a overexpression increases aneuploidy and genomic instability in FTSECs. We show that miR-181a promotes defects in nuclear structure, ruptures in the nuclear envelope, as well as mitotic and cytokinetic defects affecting proper chromosome segregation. A combination of global mRNA expression profiling and target prediction software identified the tumor suppressor retinoblastoma (RB1) as a target for miR-181a mediated FTSEC tumorigenesis. Using an RB1 3’UTR luciferase assay, along with stable expression of an shRNA against RB1 in FTSECs, we are able to show that miR-181a directly targets RB1 and that knockdown of RB1 phenocopies miR-181a mediated tumorigenesis. Taken together, our data shed light on a novel mechanism that promotes genomic instability and tumorigenesis in the early phases of HGSOC development. Our results suggest miR-181a as a promising early detection biomarker as well as a prognostic tool to guide patient treatment.

Citation Format: Matthew Knarr, Lily Kwiatkowski, Anil Nagaraj, Ronny Drapkin, Analisa DiFeo. miR-181a is a key driver of genomic instability and ovarian cancer tumorigenesis through the regulation of RB1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3557.
Abstract 3559: A crowd-control approach for castration-resistant prostate cancer therapeutics
Authors: Shukla Girish C.
Published: August 2019

Castration-resistant prostate cancer (CRPC) is defined by affecting intrinsic cellular mechanisms including dysregulated androgen signaling, aerobic glycolysis (Warburg effect), epithelial-mesenchymal transition (EMT) and dysregulation of transcription factors including androgen receptor (AR) and c-Myc. Development of CRPC post Androgen-deprivation therapy is indicated by tumor microenvironment heterogeneity, compensating cellular pathways and recurrence of AR expression leading to the failure of androgen-signaling inhibitors including Enzalutamide and Abiraterone Acetate. We have discovered that miR-644a potentiates Enzalutamide efficacy both, in vitro and in vivo prostate cancer models. In depth analyses revealed that miR-644a downregulated expression of diverse tumor microenvironment regulators including c-Myc, AR, AR co-regulators and the anti-apoptosis factors Bcl-xl and Bcl-2, EMT factors ZEB1, cdk6, and Snail. Enhanced miR-644a expression also suppressed the Warburg effect by direct down-regulating c-Myc, Akt, IGF-1R and GAPDH expression. Taken together, our data demonstrate that the adjunctive miRNA expression down-regulated several oncogenesis promoting cellular pathways and functions cooperatively with Enzalutamide. We propose that a “crowd-control” approach to pacifying aggressive tumorigenesis and metastasis supporting cellular pathways by a miRNA with a wide range of gene targets is likely to potentiate the efficacy of cancer treating drugs and potentially preventing the drug-resistance.

Note: This abstract was not presented at the meeting.

Citation Format: Girish C. Shukla. A crowd-control approach for castration-resistant prostate cancer therapeutics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3559.
Abstract 356: Novel conditionally active biologic (CAB) antibody targeting EpCAM demonstrates anti-tumor efficacy in vivo
Authors: Chang Cathy , Frey Gerhard , Sharp Leslie , Boyle William (Bill) J. , Wang Jing , Xing Charles , Liu Haizhen , Wheeler Christina , Walls Marlena , Short Jay M.
Published: August 2019

Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein mediating Ca2+-independent homotypic cell-cell adhesion during cell signaling, migration, proliferation, and differentiation. EpCAM is expressed at high levels exclusively in epithelia and epithelial-derived neoplasms, making it a suitable target for many important solid tumor types and cancer stem cells. EpCam expression on normal tissues limits its utility as target for therapeutic antibodies and ADCs due to the potential effects on normal epithelial throughout the body. Conditionally Active Biologics (CAB) technology is a proprietary platform that selects antibodies that bind to target antigens in the context of diseased tissues, but not normal tissues, by taking advantage of the unique conditions in the tumor microenvironment. Using our CAB technology, we have developed CAB antibodies to EpCAM that reversibly bind to recombinant EpCAM and EpCAM expressing cells under select conditions that are present in the tumor microenvironment but not in normal tissues. In vitro and in vivo efficacy data for several anti-EpCAM antibodies and ADCs will be presented and suggest that conditionally active EpCam ADCs generated using the CAB technology will provide drug candidates that have an increased safety margin and therapeutic index in the clinic.

Citation Format: Cathy Chang, Gerhard Frey, Leslie Sharp, William (Bill) J. Boyle, Jing Wang, Charles Xing, Haizhen Liu, Christina Wheeler, Marlena Walls, Jay M. Short. Novel conditionally active biologic (CAB) antibody targeting EpCAM demonstrates anti-tumor efficacy in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 356.
Abstract 3563: Exosomal transfer of tumor-associated macrophage derived miR-6068 promote ovarian cancer progression
Authors: Baydogan Seyda , Sheng Jianting , Kahraman Nermin , Kanlikilicer Pinar , Mokhlis Hamada Ahmed , Dilmac Sayra , Wong Stephen T. C. , Ozpolat Bulent
Published: August 2019

Ovarian cancer, especially high-grade serous ovarian cancer (HGSOC), is the deadliest gynecological cancer with 50-70% 5-year mortality rates. Every year more than 22,000 new cases of ovarian cancer and 15,000 deaths are anticipated within the United States only. Recent clinical and experimental evidence indicates that tumor-associated macrophages (TAMs), the most abundant cells in the tumor microenvironment, play a significant role in tumor growth and progression by contributing to angiogenesis, invasion, metastasis, and drug resistance, leading to poor clinical outcomes and significantly shorter patient survival in HGSOC. More than 50% of cells in the peritoneal tumor microenvironment and malign ascites consist of TAMs in ovarian cancer (OC) patients. Especially, M2 macrophages have been shown to support tumor proliferation and promote tumor progression, angiogenesis, and drug resistance. But the mechanisms of these oncogenic effects are still not clear. The goal of our study to investigate the role of TAM-derived exosomes, which are 30-100nm microvesicles released from cells and are key factors in communication between cancer cells and the tumor microenvironment. To this end, we evaluated differentially expressed miRNAs in high-grade ovarian cancer cells (OVCAR3, OVCAR 432 and OVCAR5) after treatment with exosomes-derived from TAMs (M2 phenotype) using the Affymetrix Gene Chip miRNA 4.0 microarrays. We identified several miRNAs, including miR-6068 that we validated by qPCR and found it to be significantly upregulated in both HGSOC cells and their exosomes. We demonstrated that transfection of HGSOC cells with miR-6068 significantly increased proliferation, migration and invasion capacities of the cells in vitro, suggesting that this miR-6068 act as an oncogenic miR (oncomiR). Using in silico prediction algorithms we found that miR-6068 has binding sites on the 3’-untranslated region (3’-UTR) of PTPN4 gene encoding a phosphatase and demonstrated that it miR-6068 suppresses PTPN4 expression by Western blot and qPCR. Inhibition of PTPN4 by siRNA significantly induced cell proliferation in OC cells, suggesting that PTPN4 acts as a tumor suppressor. In conclusion, our results suggest that miR-6068 has an oncogenic role in ovarian cancer progression by targeting PTPN4 and may be a novel effective therapeutic target for ovarian cancer.

Citation Format: Seyda Baydogan, Jianting Sheng, Nermin Kahraman, Pinar Kanlikilicer, Hamada Ahmed Mokhlis, Sayra Dilmac, Stephen T. C. Wong, Bulent Ozpolat. Exosomal transfer of tumor-associated macrophage derived miR-6068 promote ovarian cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3563.
Abstract 3572: SNPs in lncRNA genes are associated with non small cell lung cancer in a Chinese population
Authors: Wang Ruoyang , Qian Biyun
Published: August 2019

Background: Recent genetic studies indicate that single nucleotide polymorphisms (SNPs) in the genes encoding non-coding RNAs are associated with lung cancer susceptibility. But the comprehensive analysis of SNPs in long non-coding RNA (lncRNA) genes has not been elucidated. Methods: Based on the previous expression microarray of lncRNAs, we genotyped 17 SNPs located in 13 selected lncRNA genes by the TaqMan method in 1294 non-small cell lung cancer (NSCLC) cases and 1729 controls. Then we followed up NSCLC patients to get the survival data, and conduct logistic regression, Kaplan-Meier method and Cox hazards regression analysis to explore the associations of SNPs with NSCLC risk and overall survival (OS). Results: We identified 4 SNPs significantly associated with NSCLC risk. For rs498238, compared with CC genotype, TT genotype could decrease the risk of NSCLC (adjusted OR=0.32, 95%CI=0.11-0.92, p=0.034). As for rs16901995, TT genotype showed significantly protective effect compared with CC genotype (adjusted OR=0.76, 95%CI=0.59-0.99, p=0.040). For rs219741 dominant model, increased risk of NSCLC was evident in younger patients (age <60) compared those with wild type (adjusted OR=1.47, 95%CI=1.03-2.10, p=0.033). Rs3113503 demonstrated a controversial effect on NSCLC risk. No SNPs showed effects on overall survival of NSCL patients. Conclusion: Our findings indicated that genetic variants within lncRNA genes may predict NSCLC risk in Chinese population.

Citation Format: Ruoyang Wang, Biyun Qian. SNPs in lncRNA genes are associated with non small cell lung cancer in a Chinese population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3572.
Abstract 3577: A p53-responsive long noncoding RNA GUARDIN is essential for genome integrity
Authors: Jin Lei , Hu Wang Lai , Xu An , Wang Yu Fang , Thorne Rick F. , Zhang Xu Dong , Wu Mian
Published: August 2019

The list of long non-coding RNAs (lncRNAs) involved in the p53 pathway of the DNA damage response is rapidly expanding, but whether lncRNAs have a role in maintaining the de novo structure of DNA is unknown. Here, we demonstrate that the p53-responsive lncRNA GUARDIN is important for maintaining genomic integrity under steady-state conditions and after exposure to exogenous genotoxic stress. GUARDIN is necessary for preventing chromosome end-to-end fusion through maintaining the expression of telomeric repeat-binding factor 2 (TRF2) by sequestering microRNA-23a. Moreover, GUARDIN also sustains breast cancer 1 (BRCA1) stability by acting as an RNA scaffold to facilitate the heterodimerization of BRCA1 and BRCA1-associated RING domain protein 1 (BARD1). As such, GUARDIN silencing triggered apoptosis and senescence, enhanced cytotoxicity of additional genotoxic stress and inhibited cancer xenograft growth. Thus, GUARDIN may constitute a target for cancer treatment.

Citation Format: Lei Jin, Wang Lai Hu, An Xu, Yu Fang Wang, Rick F. Thorne, Xu Dong Zhang, Mian Wu. A p53-responsive long noncoding RNA GUARDIN is essential for genome integrity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3577.
Abstract 3581: LncRNA THAP9-AS1 promotes pancreatic cancer growth via sponging miR-484 and interacting with YAP to enhance YAP activity
Authors: Li Nan , Yang Guohua , He Zhimin , Liu Wanqing , Liu Haiying , Zheng Guopei
Published: August 2019

This study was supported by grants from the National Natural Science Foundation of China (No. 81872197, 81672616, 81401989 and 81602016); Guangdong Natural Science Funds for Distinguished Young Scholars (No.2016A030306003); Guangdong Special Support Program (No.2017TQ04R809); Science and Technology Program of Guangzhou (No.201710010100 and 201804010001).

Abnormal expression of long non-coding RNAs (lncRNAs) has been observed in various cancer types, providing probability to exploit new diagnostic markers, prognostic factors and therapeutic targets in cancer treatment. Our analysis showed lncRNA THAP9-AS1 was highly overexpressed in pancreatic cancer. Here, we aimed to investigate the roles and mechanisms of THAP9-AS1 in pancreatic cancer. We confirmed the overexpression of lncRNA THAP9-AS1 in pancreatic cancer and validated the no protein-coding potential. THAP9-AS1 promotes pancreatic cancer cells growth in vitro and in vivo. Mechanically, THAP9-AS1 exerts its effects via enhancing YAP signaling. Ectopic YAP expression disrupts the effects of THAP9-AS1 knockdown. Inversely, YAP knockdown overcomes the effects of THAP9-AS1 overexpression. THAP9-AS1acts as a competing endogenous RNA for miR-484, leading to YAP upregulation. Moreover, THAP9-AS1 binds YAP protein and inhibits the phosphorylation mediated inactivation of YAP by LATS1. Reciprocally, YAP/TEAD1 complex promotes THAP9-AS1 transcription to form a feed-forward circuit. Importantly, THAP9-AS1 level positively correlates with YAP expression in pancreatic cancer tissues. THAP9-AS1 and YAP overexpression predicts a poor outcome of pancreatic cancer patients. Our findings indicate that THAP9-AS1 plays an important role in pancreatic cancer growth via enhancing YAP signaling at two aspects. THAP9-AS1/YAP axis may serve as a potential biomarker and therapeutic target for pancreatic cancer treatment.

Note: This abstract was not presented at the meeting.

Citation Format: Nan Li, Guohua Yang, Zhimin He, Wanqing Liu, Haiying Liu, Guopei Zheng. LncRNA THAP9-AS1 promotes pancreatic cancer growth via sponging miR-484 and interacting with YAP to enhance YAP activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3581.
Abstract 3584: Role of long noncoding RNA LINC00461 in EGFR-TKIs resistant non-small cell lung cancer
Authors: Lee Ji Yun , Luu Thi-Thu-Trang , Bach Duc-Hiep , Hu Ruoci , Kim Donghwa , Lee Sang Kook
Published: August 2019

Chemo-resistance is one of the major factors limiting the therapeutic efficacy and prognosis of non-small cell lung cancer (NSCLC) patients, especially with patients who harbor epidermal growth factor receptor (EGFR)-activating mutations. Accumulating evidence suggests that aberrant expression of long noncoding RNAs (lncRNAs) is in part associated with carcinogenesis. Therefore, we tried to identify lncRNAs and their potential target genes in drug-resistant NSCLC. We evaluated lncRNAs with drug-resistant NSCLC using microarray data and in vitro assays. W found that lots of lncRNAs were differentially expressed between the drug-resistant NSCLC cells (HCC827-gef) and its parental (HCC827) cells. Among them, LINC00461 was found to be up-regulated lncRNAs, and highly correlated with the drug resistance. Taken together, the present study provides the expression profiles of chemo-resistant lncRNAs and identifies LINC00461 as a potential biomarker associated with EGFR-TKI resistance in non-small cell lung cancer cells.

This work was supported by a National Research Foundation of Korea (NRF) Grant funded by the Korean Government (MEST) (NRF-2016M3A9B6903499).

Citation Format: Ji Yun Lee, Thi-Thu-Trang Luu, Duc-Hiep Bach, Ruoci Hu, Donghwa Kim, Sang Kook Lee. Role of long noncoding RNA LINC00461 in EGFR-TKIs resistant non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3584.