bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5004: Profiling the extracellular matrix landscape of tumor microenvironments using proteomics
Authors: Naba Alexandra , Clauser Karl R. , Carr Steven A. , Hynes Richard O.
Published: August 2018

"Your cancer has spread" remains for most cancer patients synonymous with a death sentence. Indeed, the majority of cancer deaths are due to metastases: the dissemination of tumor cells from a primary tumor to distant vital organs. As of today, we do not have efficient treatments to stabilize or cure most metastatic cancers, and for patients diagnosed early, i.e., before metastases are detectable, we do not have reliable markers to predict whether their tumors will remain indolent or will become deadly. Tumor progression critically depends on a locally permissive microenvironment, including a unique tumor-associated extracellular matrix (ECM). The ECM is a complex meshwork of proteins providing architectural support to cells and conferring biomechanical properties to tissues. ECM-derived biochemical signals transduced by cell-surface receptors, such as the integrins, regulate cellular functions such as proliferation and survival, adhesion, and migration, all essential for metastasis. Importantly, clinicians have observed that more aggressive tumors present a higher ECM content. Thus, to fully understand how tumors progress and disseminate, it is essential to study the tumor ECM. To do so, we have developed a pipeline combining bioinformatics and proteomics to comprehensively characterize the ECM composition, or "matrisome," of tissues and tumors. Using this pipeline, we demonstrated that the composition of the ECM varies i) between normal tissues and tumors, ii) between poorly and highly metastatic primary tumors, and iii) between primary tumors and derived metastases. Here we present for the first time "The Tumor ECM Atlas," a compendium of ECM proteins identified by proteomics in the microenvironment of 10+ types of primary tumors and derived metastases, including breast, colorectal and lung cancers, insulinomas and multiple myelomas, which revealed the presence of a unique subset of cancer-specific and cancer type-specific ECM proteins. We believe that this atlas is a powerful tool to formulate novel hypotheses on the roles of the ECM in tumor progression and metastasis. In addition, we propose that the ECM is an underexplored reservoir of potential diagnostic and prognostic markers and therapeutic targets, and that its exploration using proteomics will pave the way for the development of novel approaches to better care for cancer patients.

Citation Format: Alexandra Naba, Karl R. Clauser, Steven A. Carr, Richard O. Hynes. Profiling the extracellular matrix landscape of tumor microenvironments using proteomics [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 5004.
Abstract 501: A RET-related microRNA, miR-153-3p, acts as a tumor suppressor in medullary thyroid carcinoma (MTC) via S6K signaling
Authors: Joo Lauren Jin Suk , Weiss Jocelyn , Gill Anthony J. , Clifton-Bligh Roderick , Brahmbhatt Himanshu , MacDiarmid Jennifer A. , Gild Matti L. , Robinson Bruce G. , Sidhu Stan B. , Zhao Jing Ting
Published: August 2018

Background

Medullary thyroid carcinoma (MTC), which comprises 3-5% of all thyroid cancers, originates from a small population of neuroendocrine calcitonin C-cells of thyroid gland. Gain-of-function mutations of the human Rearranged during Transfection (RET) proto-oncogene have been recognized as the key driver of MTC tumorigenesis. RET has been targeted by tyrosine kinase inhibitors (TKIs), however with modest efficacy. Many of the microRNAs (miRNAs) identified in human genome have been associated with cancer and have prompted interest in the establishment of miRNA-based therapeutics. However miRNA studies in MTC has been hampered due to the lack of normal C-cell tissue as a differential expression comparator.

Aims

We aimed to characterize differentially expressed miRNAs, which play fundamental roles in MTC tumorigenesis and in modulation of TKI responses.

Methods

RET of human MTC cells were silenced using TKI, Cabozantinib, or siRNA. Small RNA sequencing was performed (AGRF) to identify differentially expressed miRNAs before and after RET inhibition. MiRNA effects on cell proliferation (MTS assay), cell cycle and apoptosis (flow cytometry, Western blot) were investigated with gain-of-function studies. For in vivo study, MTC xenografts were established and miRNAs were intravenously delivered using nanoparticle delivery vehicles invented by biotechnology company - EnGeneIC Ltd.

Results

miR-153-3p was identified to be under-expressed in MTC following RET inhibition. Restoration of miR-153-3p significantly reduced cell proliferation, induced G2 cell arrest and increased cell apoptosis. Furthermore, combined treatment of miR-153-3p and Cabozantinib caused greater cell proliferation inhibition compared to individual treatment. miR-153-3p targets ribosomal protein S6 kinase B1 (RPS6KB1) and reduced phosphorylation of Bcl-2 associated death promoter (BAD) protein. In vivo delivery of miR-153-3p significantly impeded tumor growth compared to scrambled sequence. Combination of miR-153-3p delivery and oral administration of Cabozantinib resulted in further tumor stabilization effect.

Conclusion

miR-153-3p reveals a significant tumor suppressive role in MTC via S6K signaling, demonstrating its important therapeutic potential for MTC patients.

Citation Format: Lauren Jin Suk Joo, Jocelyn Weiss, Anthony J. Gill, Roderick Clifton-Bligh, Himanshu Brahmbhatt, Jennifer A. MacDiarmid, Matti L. Gild, Bruce G. Robinson, Stan B. Sidhu, Jing Ting Zhao. A RET-related microRNA, miR-153-3p, acts as a tumor suppressor in medullary thyroid carcinoma (MTC) via S6K signaling [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 501.
Abstract 5010: Advantages in using orthotopic syngeneic tumor models to evaluate immune-based approaches for cancer treatment
Authors: Maubant Sylvie , Slos Philippe , Boisferon Marc Hillairet de , Bichat Francis , Mirjolet Jean-Francois
Published: August 2018

Immune checkpoint modulators are now accepted as the fourth pillar of cancer care for both solid tumors and hematological malignancies. In addition to CTLA-4, PD-1 or PD-L1 targeting antibodies, novel immune-oncology targets are currently tested in patients but also new strategies such as bispecific T cell engagers or cell therapies including CAR-T cells. Some of them have been already approved. Prediction of preclinical models is still a long-standing debate, even more when considering modulators of the immune system. Despite significant progresses made in humanized mouse model field during the last decade, having a complete and stable immune system comprising both lymphoid and myeloid subpopulation is not yet possible. Syngeneic tumors thus remain the model of choice to interrogate and better understand the mechanisms of action of new compounds targeting immune cell interactions. In order to improve prediction and relevance of these syngeneic models, orthotopic (OT) models were developed in parallel of standard subcutaneous (SC) models. Here, comparison of regular SC versus OT engraftments are provided. Various models, such as EMT-6 (breast), MBT-2 (bladder) as well as RenCa (kidney) will be described in addition to other OT models like Hepa1-6 (liver) or PAN-02 (pancreas) models. In addition to growth characteristics using imaging for OT models, antitumor efficacy of CTLA-4 and PD-1 targeting antibodies will be detailed for both SC and OT models. One of the most interesting model is MBT-2 (bladder) for which response to CTLA-4 targeting antibody is changing depending engraftment site: significant increase in survival was evidenced for the OT model (treated to control ratio (T/C) of 285%) and only moderate efficacy was observed for the SC model (T/C = 66%). For the PD-1 targeting antibody, OT injection seemed to be the preferential site to observe increased efficacy. Attempting to correlate immune profile and response to treatment, differences in immune infiltrate between OT and SC tumors will also be presented.

Citation Format: Sylvie Maubant, Philippe Slos, Marc Hillairet de Boisferon, Francis Bichat, Jean-Francois Mirjolet. Advantages in using orthotopic syngeneic tumor models to evaluate immune-based approaches for cancer treatment [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 5010.
Abstract 5085: CXCR2 signaling in pancreatic cancer induces a tumor supporting inflammatory phenotype of the cancer-associated fibroblasts
Authors: Awaji Mohammad , Varney Michelle , Purohit Abhilasha , Batra Surinder , Singh Rakesh
Published: August 2018

Pancreatic cancer (PC) remains one of the deadliest types of solid malignancies. Making progress in dealing with PC starts by understanding the complex interaction of cellular components in the tumor microenvironment and identifying the crucial checkpoints for tumor progression and metastasis. Desmoplasia and inflammation are two major hallmarks of PC. Cancer-associated fibroblasts (CAFs) are known for inducing fibrosis in several pathologies of the pancreas including PC. Recent reports have described that CAFs can assume the fibrosis-inducing phenotype, when close or have a direct contact with tumor cells, or a tumor-promoting inflammatory phenotype, through distant paracrine signaling. Known for their role in recruiting granulocytes and inducing angiogenesis, CXC chemokines (1-3 and 5-8) contain a conserved motif of Glutamic acid-Leucine-Arginine (ELR) and signal through CXCR1/2 chemokine receptors. In PC, ELR+ chemokines produced by malignant cells and stromal cells contribute to tumor growth, angiogenesis, and tumor-supporting immunosuppression. The deletion of CXCR2 in the genetically engineered PC mouse model (KC) promoted fibrosis and tumor metastasis.

Taken together, we hypothesize that ELR+ chemokines signaling through CXCR2 may trigger the inflammatory phenotype of CAFs in PC that promotes tumor growth and progression. Utilizing unidirectional coculture studies, incubating CAFs in conditioned media of PC cells inhibited their growth, decreased the expression of the fibrotic CAFs-associated markers such as αSMA and collagen I, and increased the expression of immunosuppressive cytokines and tumor-promoting chemokines including IL-4, IL-10, IL-13, and CXCL7. Treating CAFs with exogenous ELR+ chemokines (CXCL1 and CXCL8) exhibited a similar phenotype. Furthermore, treating CAFs with exogenous ELR+ chemokines or PC cells conditioned media in the presence of pharmacological inhibitors of CXCR2 rescued this phenotype. In summary, we believe that ELR+ chemokine signaling through CXCR2 induces the inflammatory phenotype of CAFs that promotes immunosuppression, tumor growth, and tumor progression of PC.

Citation Format: Mohammad Awaji, Michelle Varney, Abhilasha Purohit, Surinder Batra, Rakesh Singh. CXCR2 signaling in pancreatic cancer induces a tumor supporting inflammatory phenotype of the cancer-associated fibroblasts [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 5085.
Abstract 5113: Spontaneous tumors in aging colonies of C57BL/6J mice
Authors: Schile Andrew , Dion Stephanie , Imai-Leonard Denise , Doty Rosalinda
Published: August 2018

The C57BL/6J inbred mouse strain is widely used to study the effects of aging. We have documented 51 spontaneous tumors (39 malignant and 12 benign) as part of a recent effort to establish large colonies of male and female C57BL/6J mice for aging research. Forty six of the tumors were diagnosed in mice submitted for histopathology from typical aging colonies maintained throughout a range of ages (6-18 months) that roughly correspond to 30 - 56 years of age in humans; the remaining cases were from much smaller cohorts aged past 18 months. Sarcomas represented the largest category of malignant tumors (20/51). There were 6 cases (5 male, 1 female) of hemangiosarcoma (4 on the neck, 1 on the liver, and 1 on the ear) and 6 multicentric histocytic sarcomas (4 male, 2 female). Four cases (1 male, 3 female) of rhabdomyosarcoma (2 on the head, 2 on the leg) were also seen. The remaining sarcoma diagnoses included one osteosarcoma (female; leg), one chondrosarcoma (male; ear), and two sarcomas not otherwise specified (both female; one leg, one lateral abdomen). Hematological malignancies constituted the second largest category (9/51), including 7 multicentric lymphomas (3 male, 4 female), a uterine lymphoma, and a multicentric plasma cell tumor (1 male). The four skin malignancies included two melanomas (male: head; female: tail/anus) and two squamous cell carcinomas (male: jaw; female: leg). Three mammary cancers were observed in females, including an adenoacanthoma and two adenocarcinomas, one of which had lung metastases. An ovarian dysgerminoma was the only germ cell tumor documented. The Harderian gland (a lacrimal duct of the eye) was the site of two carcinomas (male) and six adenomas (5 male, 1 female). Other benign tumors included a tail fibroma (male), two pulmonary adenomas (1 male, 1 female), one mammary fibroadenoma (female), a myoepithelioma of the leg (female), and a keratoacanthoma of the paw (female). Tumor incidence was very low overall, representing <0.1% of the mice present on an average week during the typical range of ages (6-18 months) in the aging colonies. Overall, the median age of diagnosis was 239 days for females and 427 days for males. These results should help to define the spectrum of tumors that develop in colonies of C57BL/6J mice that approximately correspond in age to middle-aged humans.

Citation Format: Andrew Schile, Stephanie Dion, Denise Imai-Leonard, Rosalinda Doty. Spontaneous tumors in aging colonies of C57BL/6J mice [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 5113.
Abstract 5169: Novel target molecules for treatment of cancer of unknown primary
Authors: Fujita Yoshihiko , Sakai Kazuko , Velasco Marco De , Kurata Takayasu , Hayashi Hidetoshi , Nakagawa Kazuhiko , Nishio Kazuto
Published: August 2018

We have previously been involved in a multicenter clinical study to predict the primary site of cancer of unknown primary site (CUP) for a site-specific therapy. Based on the microarray analysis of gene expression pattern, we not only predicted the primary site for each CUP patient but also extracted 44 up-regulated genes common to each CUP showing over 2.5-fold change compared to normal tissue (lymph node). To identify genes related to CUP development among these candidate genes, we performed cell-based siRNA screening and estimated to what extend A549 cells reduce the ability of migration, and found 4 genes, GRN, MIF, PRKDC, PSMB4. Konckdown of each of these genes suppressed metastasis of implanted cells from a footpad to a popliteal lymph node. Inhibitors for PRKDC (NU 7441) and PSMB4 (bortezomib) also suppressed the metastasis of parental A549 cells in vivo, suggesting the involvement of these genes in metastasis.

As there is no standard treatment for CUP, drugs targeting immune checkpoints, such as PD-1 or IDO, may reveal promising antitumor activity for CUP. Recent studies suggest that expression levels of PD-L1 (a PD-1's ligand) or IDO may be a biomarker of patient response to anti-PD-1 or anti-IDO therapy, respectively. We analyzed the expression of PD-L1 and IDO for the CUP tissues and found several patients with high expression of either or both proteins who may be most likely respond to these novel immunotherapies.

Citation Format: Yoshihiko Fujita, Kazuko Sakai, Marco De Velasco, Takayasu Kurata, Hidetoshi Hayashi, Kazuhiko Nakagawa, Kazuto Nishio. Novel target molecules for treatment of cancer of unknown primary [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 5169.
Abstract 5175: Advancing the molecular understanding of stage I colorectal cancer
Authors: Dunne Philip , Loughrey M B. , Coleman H G. , McBride R , Campbell J , Alderdice M , Redmond K L. , McArt D G. , Isella C , Leedham S , Maughan T , Lawler M , consortium S:CORT
Published: August 2018

Background: There are ~1.4 million cases of colorectal cancer (CRC) annually worldwide. Bowel cancer screening (BCS) detects cancers and high-risk adenomas earlier; previously stage I accounted for 12% of CRC, but 42% of screen-detected cancers are now stage I. This study is based on the hypothesis that within the early lesions detected by BCS there are "hopeful monsters"; a term used to describe highly aggressive tumours that are simply being caught earlier while they are still potentially curable. Colorectal tumour evolution models have proposed the “Big Bang” of tumour growth, where a single expansion in adenoma development dictates disease outcome. In line with the “hopeful monsters” theory, this model reason that some tumours are “born-to-be-bad” from the earliest point in CRC tumour evolution.

Aims: This proposal aims to develop a molecular stratifier of lethal vs non-lethal early-invasive disease based on comprehensive molecular pathological profiling, improved biological understanding and multiple tiers of validation to inform the management of CRC disease at the earliest stage. Methods: In contrast to stage II-IV CRC, there are limited stage I molecular studies, reducing opportunities to identify lethal early-disseminating tumours in patients who account for ~50% of screen-detected cancers. This study is undertaking collection and molecular profiling of a cohort of retrospective stage I tissue (n=200), enriched for patients that experienced relapsed, to identify factors associated with early-dissemination.

Results: Pathological characterisation of the stage I cohort indicated that histological factors such as fibroblast content or depth of invasive front are not associated with eventual metastatic relapse. Unsupervised analysis highlighted a detectable shift in transcriptional signalling between recurrent and non-recurrent samples. Further supervised analysis indicates that intrinsic “stem-like” factors may be more prognostic than extrinsic factors in stage I.

Conclusions: In order to find any effective treatment you have to first understand the biology underpinning disease. Given the increasing numbers of early stage patients being diagnosed as a result of BCS, and the paucity of tissue cohorts and focussed molecular studies of stage I CRC, this study aims to increase our understanding of specific factors underpinning prognosis at this early stage.

Citation Format: Philip Dunne, M B. Loughrey, H G. Coleman, R McBride, J Campbell, M Alderdice, K L. Redmond, D G. McArt, C Isella, S Leedham, T Maughan, M Lawler, S:CORT consortium. Advancing the molecular understanding of stage I colorectal 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 5175.
Abstract 520: circEPSTI1 as a prognostic marker and mediator of triple-negative breast cancer progression
Authors: Chen Bo , Huang Xiaojia , Wei Weidong , Tang Hailin , Xie Xiaoming
Published: August 2018

Purpose: Circular RNAs (circRNAs) represent a class of non-coding RNAs which play a vital role in modulating gene expression and several pathological responses. However, it is largely unknown about the expression profile and function of circRNAs in triple negative breast cancer (TNBC). In the current study, we investigated the expression profile of human circRNAs in TNBC tissues and identified circEPSTI1 (hsa_ circRNA_000479) as one circRNA significantly upregulated.

Experimental Design: We performed circular RNAs microarray assay to screen circular RNAs expression profiles of TNBC and further investigated circEPSTI1. We observed the effect of circEPSTI1 on proliferation, clonal formation and apoptosis of TNBC by knockdown of circEPSTI1 in three TNBC cell lines. According to the MREs analysis and luciferase reporter assay, we found circEPSTI1 could bind to miRNAs as a miRNA sponge and the co-target gene of miRNAs and performed xenografts in mice to confirm our findings. We evaluated circEPSTI1 levels of 240 TNBC patients by ISH.

Results: We found that knockdown of circEPSTI1 could inhibit TNBC cell proliferation and induce apoptosis. In vitro and in vivo experiments implicated that circEPSTI1 could bind to miR-4753 and miR-6809 as miRNAs sponge to regulate BCL11A expression and affect TNBC proliferation and apoptosis. High level of circEPSTI1 was correlate with shorter survival in TNBC patients.

Conclusions: The circEPSTI1-miR-4753/6809-BCL11A axes can affect the proliferation and apoptosis of triple negative breast cancer through the mechanism of competing endogenous RNAs. Meanwhile, our results provide circEPSTI1 as an independent prognostic marker for survival in patients with TNBC.

Citation Format: Bo Chen, Xiaojia Huang, Weidong Wei, Hailin Tang, Xiaoming Xie. circEPSTI1 as a prognostic marker and mediator of triple-negative breast cancer progression [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 520.
Abstract 5348: Single-cell analysis of mutational heterogeneity in acute myeloid leukemia tumors with high-throughput droplet microfluidics
Authors: Eastburn Dennis J. , Pellegrino Maurizio , Sciambi Adam , Treusch Sebastian , Xu Liwen , Durruthy-Durruthy Robert , Gokhale Kaustubh , Jacob Jose , Chen Tina X. , Oldham William , Matthews Jairo , Kantarjian Hagop , Futreal Andrew , Patel Keyur , Jones Keith W. , Zehnder James L. , Takahashi Koichi
Published: August 2018

To make possible the routine characterization of genetic diversity within cancer cell populations, we developed a novel two-step microfluidic droplet workflow that enables efficient and massively-parallel single-cell PCR-based genomic barcoding for single-cell DNA sequencing applications. We demonstrate that the two-step microfluidic approach is required for robust DNA amplification on thousands of individual cells per run with high coverage uniformity and low allelic dropout of targeted genomic loci. To apply our single-cell sequencing technology to human tumor samples, we developed a targeted panel to partially sequence 26 genes frequently mutated in acute myeloid leukemia (AML) including TP53, DNMT3A, FLT3, NPM1, NRAS, IDH1 and IDH2. Using this panel, we were able to sensitively identify SNV and indel-defined clones within AML samples and assess their distribution at the time of diagnosis, remission and relapse. We also used single cell SNVs to monitor host and donor cell populations during bone marrow transplantation (BMT), which allowed us to accurately evaluate engraftment and disease relapse. Collectively, our single-cell data indicates that clonal populations inferred from VAFs obtained from bulk sequencing data may not fully resolve the heterogeneity within tumors; moreover, the single-cell nature of our approach enabled the unambiguous identification of multiple co-occurring mutations within subclones that is not possible with bulk measurements. Collectively, our results show a greater degree of heterogeneity in AML tumor samples than is commonly appreciated with traditional sequencing paradigms and demonstrate the value of single-cell analysis for AML.

Citation Format: Dennis J. Eastburn, Maurizio Pellegrino, Adam Sciambi, Sebastian Treusch, Liwen Xu, Robert Durruthy-Durruthy, Kaustubh Gokhale, Jose Jacob, Tina X. Chen, William Oldham, Jairo Matthews, Hagop Kantarjian, Andrew Futreal, Keyur Patel, Keith W. Jones, James L. Zehnder, Koichi Takahashi. Single-cell analysis of mutational heterogeneity in acute myeloid leukemia tumors with high-throughput droplet microfluidics [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 5348.
Abstract 5359: Regulatory germline variants in 10,389 adult cancers
Authors: Huang Kuan-lin , Weerasinghe Amila , Wu Yige , Liang Wen-wei , Mashl R. Jay , Reynolds Sheila , Houlahan Kathleen E. , Oak Ninad , Atlas The Cancer Genome , Lazar Alexander J. , Wendel Michael C. , Khurana Ekta , Plon Sharon , Chen Feng , Gerstein Mark , Shmulevich Ilya , Ding Li
Published: August 2018

Previous studies of rare germline variants in cancer has largely been limited to the coding regions of known predisposition genes. The TCGA PanCanAtlas Germline Working Group is analyzing germline predisposing variants of 10,389 cases in 33 cancer types. We deployed more than 121,000 virtual machines running for over 600,000 hours on the ISB Cancer Genome Cloud to conduct massively parallel variant calling and analyses, and the resulting data are shared with scientists across institutions worldwide. Carriers of the functional regulatory variants add on to the 8.9% of cases carrying known pathogenic variants. Burden analyses reveal enrichment of rare variants in the 3'UTR region of NHP2 and POLH. Further, we observed variants aggregating in conserved regions of selected microRNA families that are also affected by somatic mutations, including mir-17 and mir-29. We nominate regulatory variants by using GWAVA and FunSeq2 corroborated with their enrichment in cancer. The prioritized variants are then further evaluated by further co-occurrence of two-hit events and expression changes in their respective tumor samples. Finally, we examine ancestries, familial history and age at onset for carriers of these variants. Overall, we aim to discover and establish the role of regulatory germline variants in oncogenesis.

Citation Format: Kuan-lin Huang, Amila Weerasinghe, Yige Wu, Wen-wei Liang, R. Jay Mashl, Sheila Reynolds, Kathleen E. Houlahan, Ninad Oak, The Cancer Genome Atlas, Alexander J. Lazar, Michael C. Wendel, Ekta Khurana, Sharon Plon, Feng Chen, Mark Gerstein, Ilya Shmulevich, Li Ding. Regulatory germline variants in 10,389 adult cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5359.
Abstract 5461: B7-H3 regulates NF-kB signaling in cancer cells by modulating the degradation and re-synthesis of IkBα
Authors: Glazer Sarah , Gammon Seth , Piwnica-Worms David
Published: August 2018

B7-H3, a B7 homologue, is highly expressed on tumors and tumor vasculature. Although its role as a T cell inhibitor or co-stimulator is still under investigation, high expression of membranous B7-H3 in the tumor compartment is correlated with poor overall survival. NF-kB activation, standing at the intersection between inflammation and cancer, can lead to both the production of cytokines that engage the innate immune system, as well as pro-proliferative signals in cancer. B7-H3 has been shown to enhance and suppress inflammatory signaling by modulating NF-kB. However, the mechanism by which this occurs in cancer cells remains unknown. In this study, we aim to understand how B7-H3 modulates the NF-kB pathway in order to regulate incoming inflammatory signals in cancer cells. The human colon cancer cell line, HCT116, was stably transfected with a transcriptionally-coupled post-translational NF-kB luciferase reporter comprising of the inhibitor of NF-kB, IkBα, fused to firefly luciferase and placed downstream of an NF-kB responsive promoter, kB5. This construct allows for the real-time readout of the degradation of IkBα and NF-kB driven transcriptional re-synthesis of IkBα in response to a stimulus. The reporter cell line was treated with two stimulants, TNFα and flagellin, that activate NF-kB through different pathways and to different degrees with or without treatment with human B7-H3. Treatment of the reporter cell line with B7-H3 alone did not induce IkBα degradation or re-synthesis. However, stimulation of the reporter cell line with TNFα or flagellin in combination with B7-H3 attenuated the degradation of IkBα and its re-synthesis compared to stimulation with TNFα or flagellin alone. Maximal re-synthesis of IkBα when stimulated with TNFα was 8.25 ± 1.51 fold (S.D.) and significantly increased compared to cells additionally treated with B7-H3 (5.83 ± 1.23 fold, p = .01). In addition, peak degradation time of IkBα when cells were stimulated by TNFα was 45 ± 5 min (S.D.), and significantly shorter than when cells were also treated with B7-H3 (61.67 ± 2.89 min, p = .01). Similar trends were observed when cells were treated with both flagellin and B7-H3 compared to stimulation with flagellin alone. These studies show that B7-H3 suppresses the response of colon cancer cells to incoming inflammatory stimuli and attenuates NF-kB signaling by tempering the kinetics and dynamics of IkBα.

Citation Format: Sarah Glazer, Seth Gammon, David Piwnica-Worms. B7-H3 regulates NF-kB signaling in cancer cells by modulating the degradation and re-synthesis of IkBα [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 5461.
Abstract 5481: Targeting the kynurenine pathway as a novel metabolic treatment for head and neck cancer
Authors: Subramanian Chitra , Rajendiran Thekkelnaycke M. , Soni Tanu , Cohen Mark S.
Published: August 2018

Introduction: Head and neck squamous cell carcinoma (HNSCC) still claims 330,000 lives globally each year, indicating a critical need for novel therapies. Recently, we identified high levels of kynurenine (K) in HNSCC patients through global metabolic profiling of HNSCC tumor tissues via mass spectrometry (MS). Indoleamine-2,3-dioxygenase (IDO) and tryptophan-2,3- dioxygenase (TDO) are enzymes that degrade tryptophan to K. Elevated K allows tumor cells to evade immune response more easily, promoting cancer progression, while increased IDO levels are associated with poorer prognosis. We hypothesize that restoring immune response to tumors by blocking IDO1 will be an effective therapeutic strategy for sensitizing HNSCC.

Methods: Validated human HNSCC cell lines were grown in 2D culture. Kynurenine levels were measured by MS. IDO1, IDO2 and TDO levels in tissue samples were measured by Western blot (WB) analysis. siRNAs for IDO1, IDO2, and TDO were used to knock down tryptophan pathway enzymes (non-targeting siRNA controls). Knockdown efficiency was evaluated by immunocytochemistry. After knockdown of IDO1, RNA was isolated, reverse transcribed and cancer associated pathways were analyzed using RT2 cancer pathway profiler (data analyzed using GeneGlobe; Qiagen).

Results: Evaluation of the expression levels of enzymes that play a critical role in kynurenine pathway by WB analysis indicated upregulation of IDO1, IDO2 and TOD in both primary and metastatic tumor samples. siRNA-mediated knockdown of these enzymes indicated that only IDO1 knockdown was able to significantly reduce the levels of kynurenine by more than 80% (p<0.001), whereas knockdown of IDO2 and TDO reduced the kynurenine levels only by 10-25% (p>0.05%). Regulation of IDO1 function evaluated by the RT2 cancer pathway finder indicated downregulation of several genes involved in angiogenesis (ANGPT1, MCP-1, FGF2, TEK, VEGF, SERPINF1), pro-survival (c-IAP2 and XIAP), EMT transition (CDH2, OCLN and SNAI1), and DNA repair (LIG4, POLB, ERCC3 and DDB2), whereas genes involved in glycolysis (ACLY, G6PD,COX5A,LPL and PFKL), DNA damage (GADD34, GADD45G etc) and apoptosis (CASP7, CASP9, BCL2L11) were upregulated.

Conclusion: Kynurenine is highly upregulated in HNSCC with checkpoint inhibition of IDO1 leading to increased apoptosis in vitro as well as increases in several glycolytic enzymes. Further translational analysis of glycolytic influences on the tryptophan pathway and immune checkpoint inhibition of IDO1 will provide mechanistic support for development of novel metabolic therapies for HNSCC.

Citation Format: Chitra Subramanian, Thekkelnaycke M. Rajendiran, Tanu Soni, Mark S. Cohen. Targeting the kynurenine pathway as a novel metabolic treatment for head and neck 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 5481.
Abstract 5561: High-affinity antisense oligonucleotides targeting Foxp3 inhibit immunosuppressive function of regulatory T-cells and produce antitumor effects in syngeneic tumor models
Authors: Sinclair Charles , Revenko Alexey S. , Peter Alison , Johnson Robert B. , Hettrick Lisa A. , Taylor Molly , Staniszewska Anna , Hughes Adina , Sandin Lisa , Klein Stephanie , Watt Andrew , Barry Simon , Monia Brett T. , Lyne Paul , Macleod A Robert , Edbrooke Mark
Published: August 2018

Regulatory T-cells (Treg) contribute to cancer progression by suppressing anti-tumor immuity. Tregs specifically require expression of the lineage defining transcription factor Foxp3 for their development and function, but this protein cannot be targeted with conventional small molecule or biologic drugs. We employed next-generation antisense inhibitors (Gen 2.5 cEt-modified ASOs) in an attempt to selectively inhibit Foxp3 expression in mouse Treg cells, and evaluated consequences of ASO-mediated Foxp3 knock-down in vitro and in vivo. Mouse Foxp3-specific ASOs promoted potent dose-dependent reductions in Foxp3 mRNA and protein in vitro, without the use of transfection reagents. Foxp3 knockdown also resulted in loss of immunosuppressive markers and conferred Tregs with a reduced immunosuppressive capacity. Whilst genetic ablation of FOXP3 leads to manifestation of autoimmunity due to a complete loss of Treg function, we observed that the systemic delivery of unformulated mouse Foxp3 ASOs to WT mice resulted in Foxp3 knockdown in vivo, but spared mice from an autoimmune phenotype. Nevertheless, when syngeneic tumor-bearing mice were treated with mouse Foxp3 ASOs tumor growth was significantly attenuated, with a fraction of animals (25%-50%) achieving complete regressions. Anti-tumor activity of mouse Foxp3 ASOs was associated with immunophenotypic changes consistent with an increased anti-tumor immune response. Overall these data demonstrate the therapeutic capacity of mouse ASO to directly target regulatory T-cells, and suggesting that targeting of Foxp3 represents an attractive opportunity in cancer immunotherapy.

Citation Format: Charles Sinclair, Alexey S. Revenko, Alison Peter, Robert B. Johnson, Lisa A. Hettrick, Molly Taylor, Anna Staniszewska, Adina Hughes, Lisa Sandin, Stephanie Klein, Andrew Watt, Simon Barry, Brett T. Monia, Paul Lyne, A Robert Macleod, Mark Edbrooke. High-affinity antisense oligonucleotides targeting Foxp3 inhibit immunosuppressive function of regulatory T-cells and produce antitumor effects in syngeneic tumor models [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 5561.
Abstract 5586: Circulating tumor cells as the prognostic marker for non-remission and disease-specific mortality of patients with thyroid cancer
Authors: Tseng Ching-Ping , Lin Jen-Der , Liou Miaw-Jene , Hung Wei-Shan , Hsu Hsueh-Ling , Leong Kong-Kit , Chen Yu-Ting , Cheng Ju-Chien
Published: August 2018

Patients with thyroid cancer usually have good prognosis. However, 15-20% of patients ultimately develop recurrence and disease-related death. In this study, we investigate whether the number of circulating tumor cells (CTCs) expressing either epithelial cell adhesion molecule (EpCAM), podoplanin (PDPN, a marker related to poor prognosis in patients with thyroid cancer), or thyroid-stimulating hormone receptor (TSHR, a marker for thyroid cells) is a prognostic marker for non-remission and disease-specific mortality (DSM) of patients with thyroid cancer. Blood samples were collected from patients (n = 128) after thyroidectomy or radioactive iodide therapy. After enrichment of CTCs by a negative selection PowerMag system, enumeration and subtyping of the CD45-depleted cells were performed by immunofluorescence staining using the antibodies aginst EpCAM, TSHR, and PDPN, respectively. Our data revealed that the number of EpCAM+-CTCs (p < 0.001) and PDPN+-CTCs (p < 0.001), and TSHR+-CTCs (p < 0.001) for patients in the non-remission group (n = 43) was significantly higher when compared to the remission group (n = 85). At the cutoff of 40, 14, and 47 cells/mL for EpCAM+-CTCs, TSHR+-CTCs, and PDPN+-CTCs, the accuracy of the assay was equivalent to 80.4%, 76.6%, and 77.3%, respectively. On the other hand, the number of EpCAM+-CTCs (p < 0.001), PDPN+-CTCs (p = 0.013), and TSHR+-CTCs (p < 0.001) for patients in the DSM group (n = 17) was significantly higher when compared to the patients who survived (n = 111). At the cutoff of 27, 25, and 9 cells/mL for EpCAM+-CTCs, TSHR+-CTCs, and PDPN+-CTCs, the accuracy of the assay was equivalent to 69.5%, 67.2%, and 68.5%, respectively. These data together indicate that CTC testing is worthy to be considered as a routine clinical test to benefit clinical care of patients with thyroid cancer.

Citation Format: Ching-Ping Tseng, Jen-Der Lin, Miaw-Jene Liou, Wei-Shan Hung, Hsueh-Ling Hsu, Kong-Kit Leong, Yu-Ting Chen, Ju-Chien Cheng. Circulating tumor cells as the prognostic marker for non-remission and disease-specific mortality of patients with thyroid 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 5586.
Abstract 5626: Preclinical characterization of Sym022, a novel anti-LAG3 antibody
Authors: Grandal Michael M. , Melander Maria C. , Bhatia Vikram K. , Gjetting Torben , Lindsted Trine , Fröhlich Camilla , Lantto Johan , Horak Ivan D. , Kragh Michael , Kofoed Klaus , Pedersen Mikkel W.
Published: August 2018

Immunotherapy has become a major focus of research in oncology and blockade of immune checkpoints such as cytotoxic T-Lymphocyte associated protein 4 (CTLA4) and programmed cell death protein 1 (PD1) has been some of the most successful immunotherapies. Lymphocyte-activation gene 3 (LAG3) belongs to the second-generation immune modulatory targets. LAG3 is expressed by activated T-cells and tumor infiltrating lymphocytes (TILs) and negatively regulates T-cell activity upon ligand engagement. LAG3 binds major histocompatibility complex class II (MHCII) molecules found on the surface of antigen presenting cells and tumor cells. Sym022 is a Fc-inert human monoclonal antibody targeting LAG3. Sym022 binds to human and cynomolgus monkey LAG3 with high affinity and blocks the interaction between LAG3 and MHCII molecules. Functionally, Sym022 increases cytokine production by T-cells in vitro and tumor growth inhibition in vivo. Mechanistically, Sym022 not only blocks ligand binding, but also decreases total LAG3 surface levels through internalization and/or shedding.

Citation Format: Michael M. Grandal, Maria C. Melander, Vikram K. Bhatia, Torben Gjetting, Trine Lindsted, Camilla Fröhlich, Johan Lantto, Ivan D. Horak, Michael Kragh, Klaus Kofoed, Mikkel W. Pedersen. Preclinical characterization of Sym022, a novel anti-LAG3 antibody [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 5626.
Abstract 5748: Downregulation of glycine decarboxylase renders ovarian cancer cells less proliferative and more chemoresistant
Authors: Shin So-Jin , Chung Hyewon , Kim Jin Young , Kim Hyera , Cho Chi-Heum , Ha Eunyoung
Published: August 2018

Glycine decarboxylase (GLDC) is a very recently identified metabolic oncogene that links glycine metabolism with tumorigenesis. We designed this study to determine the role of GLDC in chemoresistance in ovarian cancer cells. First, we found GLDC is downregulated in paclitaxel-resistant ovarian cancer cells (OVCAR3/PTX). Prompted by decreased glucose transport in OVCAR3/PTX cells, we transplanted OVCAR3 and OVCAR3/PTX onto the back of the NOD-scid IL2Rgammanullmice and found that the OVCAR3/PTX tumor grew less rapidly than paclitaxel-sensitive ovarian cancer cells (OVCAR3). We also found that GLDC is downregulated in OVCAR3/PTX tumor. In vitro study revealed that the inhibitory effect of siGLDC on proliferation was significantly greater in OVCAR3 than in OVCAR3/PTX. We also found that knockdown of GLDC in OVCAR3 cells rendered ovarian cancer cells PTX-resistant. Finally, we determined GLDC expressions in patients with chemosensitive and chemoresistant serous-type ovarian cancer and found that GLDC expression was markedly downregulated in chemoresistant cancer patients. In summary, these results suggest that GLDC is associated with ovarian cancer chemoresistance.

Citation Format: So-Jin Shin, Hyewon Chung, Jin Young Kim, Hyera Kim, Chi-Heum Cho, Eunyoung Ha. Downregulation of glycine decarboxylase renders ovarian cancer cells less proliferative and more chemoresistant [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 5748.
Abstract 5802: Efficacy of PI3Kβ inhibitor AZD8186 in PTEN-deficient triple-negative breast cancer
Authors: Owusu-Brackett Nicci , Zhao Ming , Akcakanat Argun , Evans Kurt W. , Yuca Erkan , Meric-Bernstam Funda
Published: August 2018

Background: Phosphatase and tensin homologue (PTEN) loss is a frequently disrupted tumor suppressor in cancer. PTEN serves as a negative regulator of the phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR pathway to promote balanced cell proliferation, survival and differentiation. Consequently, loss of PTEN function increases cell proliferation, decreases cell death and contributes to tumor initiation. PTEN loss occurs across a variety of cancer subtypes, and these PTEN deficient tumors are dependent on PI3Kβ activity. As a result, therapeutic approaches that focus on the inhibition of PI3Kβ isoform in PTEN-null cells are of interest. We evaluate the efficacy of PI3Kβ inhibitors focused on triple negative breast cancers with loss of PTEN function. Methods: We studied the effects of p110β inhibitor AZD8186 in a panel of ten TNBC cell lines, which included four PTEN-mutant cell lines. Three of four cell lines were confirmed to have PTEN loss on western blot. In vitro, cell proliferation assays were performed to determine the half maximal inhibitory concentration (IC50) after 3 days of treatment and to test the effects in combination with chemotherapy. We confirmed the sensitivity of the PTEN-null cell lines to AZD8186 with colony formation assays. Western blot analysis was performed to assess PTEN expression and PI3K pathway activation in MDA-MB-436 and MDA-MB-468 cell lines, and to evaluate effects of AZD8186 on PI3K signaling. Results: Cell lines with PTEN loss were significantly more sensitive to AZD8186 in vitro (p= 0.008). AZD8186 inhibited PI3K signaling. Western blot revealed decreased activation of pAKT, pGSK3β, pPRAS40 and pS6. Activation of other pathways were evaluated with no activation of the MAPK/ERK or MEK pathways appreciated. AZD8186 treatment resulted in increased apoptosis but did not have a significant effect on cell-cycle progression in PTEN-deficient cell lines. AZD8186 was not synergistic with eribulin, paclitaxel or carboplatin; combinations with novel targeted therapies in vitro and in vivo are ongoing. Conclusion: AZD8186 had single agent efficacy in PTEN-deficient triple negative breast cancer cell lines in vitro. Further study is needed to identify rationale combinations and in vivo efficacy.

Citation Format: Nicci Owusu-Brackett, Ming Zhao, Argun Akcakanat, Kurt W. Evans, Erkan Yuca, Funda Meric-Bernstam. Efficacy of PI3Kβ inhibitor AZD8186 in PTEN-deficient triple-negative breast 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 5802.
Abstract 5816: Inhibition of Hsp90 sensitizes pancreatic cancer in vitro and in vivo to chemo-radiotherapy
Authors: Nagaraju Ganji Purnachandra , El-Rayes Bassel F.
Published: August 2018

Purpose: Important signaling pathways contributing to proliferation, development, metastasis and resistance to chemo-radiotherapy in pancreatic adenocarcinoma (PDAC) includes the PI3K/AKT/HIF-1α/STAT-3 pathway. The molecular chaperone heat shock protein 90 (HSP90) controls the activity, stability, turnover, and trafficking of many molecules involved in these major signaling pathways. Functional inhibition of HSP90 may inhibit these pathways simultaneously. We evaluated the efficacy of HSP90 functional inhibitor ganetespib in combination with chemo-radiotherapy against PDAC in vitro and in vivo.

Methods and Materials: Cell proliferation (Br-dU), colony formation (clonogenic assay), migration (spheroid), and Western blot analyses were carried out in HPAC and PANC-1 PDAC cell lines to determine the effect of ganetespib alone or in combination with 5-FU+ 2Gy XRT. Tumor regression study of PDAC with ganetespib alone and in combination with chemo-radiation was analyzed in xenograft mouse models.

Results: At the clinically achievable dose of 50 nM, ganetespib inhibited PDAC cells proliferation (p<0.001), potentiated 5-FU chemotherapy and 2Gy of ionizing radiation (p<0.001), and decreased the levels of survival molecules such as PI3K, p-AKT, and p-ERK in vitro. Additionally, ganetespib indeed potentiates the effects of chemo-radiation in both cell lines: significant decrease in colony formation as well as decrease in migration. Combinations of ganetespib and chemo-radiation therapy demonstrated synergistic ability in human xenograft mouse models (p<0.001).

Conclusion: Ganetespib potentiates the effects of chemo-radiation in preclinical studies. Evaluating HSP90 inhibitors in PDAC patients is a rational approach in assessing the fatal PDAC.

Citation Format: Ganji Purnachandra Nagaraju, Bassel F. El-Rayes. Inhibition of Hsp90 sensitizes pancreatic cancer in vitro and in vivo to chemo-radiotherapy [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 5816.