bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3549: C-MYC sensitizes GBM with primitive features to glutamine metabolism disruption
Authors: Poore Brad Andrew , Taylor Isabella , Rubens Jeffrey , Hanaford Allison , Maxwell Micah , Eberhart Charles , Raabe Eric
Published: July 2017

Glioblastoma (GBM) is among the most lethal of known human cancers, with a median survival of less than 15 months. The highly infiltrative nature and genetic heterogeneity of GBM renders treatment difficult. Therefore, better and more targeted therapies are needed for patients with GBM. There is a new WHO subset of GBM that contains primitive neuronal components (GBM-PNC). These tumors can arise from a histologically classic GBM, and often the GBM-PNC portions of the tumor contain C-MYC or N-MYC amplifications. High MYC expression is known to alter cellular metabolism, increasing reliance on glutamine, which may create opportunities for therapeutic intervention. We hypothesized that depriving GBM-PNC cells of glutamine using metabolic inhibitors would suppress growth and tumorigenicity. To create genetically appropriate GBM-PNC models, we derived cortex (CTX) human neural stem cells and transformed them through lentiviral expression of mutant p53, constitutively-active AKT and hTERT. Transformed neurospheres were then lentivirally transduced with either C-MYC or BMI1. These models formed aggressive tumors in mice and recapitulated the histological features of GBM with expression of NESTIN, GFAP, and MAP2. When treated with the glutamine metabolic inhibitors DON or Acivicin, transformed neurospheres that expressed C-MYC had decreased cellular proliferation (BrdU incorporation, P<0.05) and increased apoptosis (cleaved caspase-3 expression, P<0.05) while neurospheres expressing BMI1 had no statistically significant changes. Additionally, cells overexpressing C-MYC had an increased expression of glutaminase, an essential enzyme for glutamine incorporation into the TCA cycle. We hypothesize that C-MYC expression sensitizes GBM-PNET cancer cell lines to DON or Acivicin inhibition through greater reliance upon glutamine for its TCA cycle intermediates. In addition C-MYC upregulates multiple glutamine transporters, which may also be used to transport DON and/or Acivicin inside the cell, conferring increased sensitivity. After further validating glutamine metabolic inhibitors in orthotopic xenografts and investigating the mechanism of action by metabolomics, we anticipate that metabolic inhibitors may have a therapeutic role in GBM-PNC.

Citation Format: Brad Andrew Poore, Isabella Taylor, Jeffrey Rubens, Allison Hanaford, Micah Maxwell, Charles Eberhart, Eric Raabe. C-MYC sensitizes GBM with primitive features to glutamine metabolism disruption [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3549. doi:10.1158/1538-7445.AM2017-3549
Abstract 3553: Mutant KRas-mediated AKT2 activation supports lung cancer growth by activating complex II-driven mitochondrial metabolism
Authors: Park Kwan-Hee , Lee Ho-Young
Published: July 2017

Non-small cell lung cancer (NSCLC), the most common type of lung cancer (comprising 80-85 %), is a leading cause of cancer-related death worldwide. KRAS mutation is s major driver oncogene in NSCLC. Constitutive activation of K-Ras via oncogenic mutations plays a key role in development and progression of NSCLC, and is an important target for drug development. Indeed, drugs that target RAF-MAPK and PI3K, the well-established downstream pathways of K-Ras, are being evaluated in the clinic. However, to date, no effective treatments specifically targeting K-Ras have reached the clinic. A new direction for anti-Ras therapies targeting Ras-mediated altered cellular metabolism is implied by recent studies. Hence, as an attempt to investigate novel therapeutic targets to suppress KRAS-driven NSCLC, we assessed metabolic changes in NSCLC cell line and mouse model systems with oncogenic K-Ras mutations. We show that activation of Ras causes metabolic alterations, leading to aerobic glycolysis. Interestingly, despite the changes in the glycolytic pathway, NSCLC cell line and mouse models carrying K-Ras mutation revealed an elevated mitochondrial membrane potential, superoxide production as well as maximal oxygen consumption rate. Moreover, blockade of mitochondrial respiration effectively reduced mutant KRas-driven neoplastic phenotypes in NSCLC cell lines vitro and lung tumor formation in vivo. In contrast, blockade of glycolysis was not effective in the mutant KRas-driven tumorigenic activities. Mechanistically, mutant KRas-mediated activation of Akt2 led to increased expression of succinate dehydrogenase subunit A (SDHA) through increased protein stability by blocking proteasome-mediated degradation, resulting in mitochondrial complex II-dependent respiration. Our study suggests that induction of mitochondrial complex II-dependent respiration is an important mechanism by which oncogenic mutant KRas induces metabolic changes and ROS stress in NSCLC cells, contributing to lung carcinogenesis. Our findings elucidate potential metabolic targets that may provide novel therapeutic strategy for treating NSCLC with oncogenic KRAS mutations.

Citation Format: Kwan-Hee Park, Ho-Young Lee. Mutant KRas-mediated AKT2 activation supports lung cancer growth by activating complex II-driven mitochondrial metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3553. doi:10.1158/1538-7445.AM2017-3553
Abstract 3555: Lipid droplets, a novel metabolic target in glioblastoma
Authors: Geng Feng , Guo Jeffrey , Cheng Xiang , Wu Xiaoning , Cheng Chunming , Chakravarti Arnab , Guo Deliang
Published: July 2017

Lipid metabolism reprogramming is a novel feature of malignancies. Identifying the vulnerability of lipid alteration in tumor cells provides a new window to treat cancer. Here, we unraveled that glioblastoma (GBM) patients containing high levels of lipid droplets (LDs), special organelles storing cholesterol esters and triglycerides in cells, presented poor survival. We found that LDs largely formed in tumor tissues from GBM patients, but rarely in low grade of gliomas and normal brain tissues, demonstrating that LDs might serve as a novel diagnostic biomarker and prognostic marker for GBM. We further revealed that sterol O-acyltransferase 1 (SOAT1), the key enzyme controlling cholesterol esterification and LD formation, is highly expressed and correlated with LD prevalence in tumor tissues from GBM patients. Genetic or pharmacologic inhibition SOAT1 using avasimibe, a phase III clinical trial drug in human atherosclerosis patients, significantly suppressed GBM growth and prolongs survival in xenograft models. Moreover, we found that inhibiting SOAT1 suppressed sterol regulatory element-binding protein-1 (SREBP-1), a key transcription factor playing a central role in lipid synthesis, underlying the molecular mechanism of targeting SOAT1-mediated GBM growth suppression. In summary, our data demonstrate that LDs/cholesterol esters uniquely formed in tumor tissues provides an ideal target to specifically inhibit GBM cells whereas sparing normal brain tissues, and reveal that blocking cholesterol storage via targeting SOAT1 represents a promising strategy for GBM treatment.

Citation Format: Feng Geng, Jeffrey Guo, Xiang Cheng, Xiaoning Wu, Chunming Cheng, Arnab Chakravarti, Deliang Guo. Lipid droplets, a novel metabolic target in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3555. doi:10.1158/1538-7445.AM2017-3555
Abstract 3567: Sequencing the human TCRβ repertoire on the Ion S5™ System
Authors: Topacio-Hall Denise S. , Looney Tim , Sun Yongming , Miller Lauren , Linch Elizabeth , Lowman Geoffrey , Lin Lifeng , Andersen Mark , Hyland Fiona , Mongan Ann
Published: July 2017

Next-generation Sequencing (NGS) is proving an important tool in increasing understanding of the human immune system, and thereby cancer immunology. αβ-T cells are the primary constituents of human cell-mediated adaptive immunity. The antigen specificity of each αβ-T cell is encoded in the 500-600 bp transcript encompassing the variable portion of the rearranged TCRα and TCRβ subunits, which can be read via NGS in a process termed repertoire sequencing. Until now, the main challenge the field faces is the lack of a technology that can provide a contiguous read of 600 bp to minimize the complexity of designing bias-prone primers and informatics challenges of stitching short reads. Here we leverage the long read capability of Ion 530™ chip to comprehensively sequence all three CDR domains of the TCRβ chain. The Ion 530™ chip offers greater than 15 M productive reads, allowing a multiplex of 2-4 samples with sufficient coverage for most repertoire profiling studies. Initial testing with Leukocyte total RNA demonstrates that this multiplex PCR assay produced repertoires that were much more similar to data derived from 5’RACE protocol than the commonly used BIOMED2 primer set. This result suggested that the use of long reads minimizes bias by allowing targeting of less variable regions. To further assess the performance of the assay, we designed a model system of 30 plasmid controls containing common human T-cell CDR3 sequences. Each plasmid was amplified individually and sequenced to confirm the detection of a single clonal population. Analytical sensitivity of the assay and accuracy of the accompanied analysis solution were further evaluated by spiking in plasmid concentrations from 10 pg to 0.0001 pg (5 million to 50 copies) in a background of 100 ng cDNA reverse transcribed from leukocyte total RNA. Results showed the assay offers linearity over 5 orders of magnitude of decreasing input concentration. In summary, we have demonstrated a NGS workflow for TCRβ sequencing that offers multiplex flexibility on Ion S5 with sample to answer in less than 48 hours. For Research Use Only. Not for use in diagnostic procedures.

Citation Format: Denise S. Topacio-Hall, Tim Looney, Yongming Sun, Lauren Miller, Elizabeth Linch, Geoffrey Lowman, Lifeng Lin, Mark Andersen, Fiona Hyland, Ann Mongan. Sequencing the human TCRβ repertoire on the Ion S5™ System [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3567. doi:10.1158/1538-7445.AM2017-3567
Abstract 357: NKX2-1 regulates SMAD and non-SMAD pathways in thyroid stem cells
Authors: Iwadate Manabu , Takizawa Yoshinori , Kimura Shioko
Published: July 2017

NKX2-1 plays a critical role in thyroid differentiation. Recently, we established Side Population cells-derived Thyroid cell line (SPTL) from mouse thyroid Side Population cells. In SPTL cells, NKX2-1, PAX8 and E-cadherin were not expressed when examined by western blotting. Immunocytochemistry revealed that only 1% of SPTL cells were NKX2-1 positive. When GFP-SPTL cells were directly injected into mouse thyroid, a few GFP-SPTL cells with weak expression of NKX2-1 were found in part of thyroid follicle, suggesting that SPTL cells have potential of thyroid stem cells. Comprehensive gene expression analysis indicated that TGF-beta signaling pathway is critical for SPTL cells. TGF-beta signaling and NKX2-1 appear to play an important role in thyroid stem cells and thyroid differentiation. Here we show that NKX2-1 regulates both SMAD and non-SMAD pathways in SPTL cells. To examine the role of NKX2-1 in SPTL cells, we produced SPTL cells having doxycycline-inducible Nkx2-1 expression (named SPTL(Nkx2-1on)). In SPTL(Nkx2-1on) cells, doxycycline induced NKX2-1 as expected, while PAX8 and E-cadherin expression was not. Upon overexpression of NKX2-1 in SPTL(Nkx2-1on) cells, TGF-beta-induced phospho-SMAD3 expression was inhibited. Additionally, NKX2-1 enhanced the expression of phospho-AKT. In GFP-myrAKT(constitutively active AKT)-overexpressing SPTL cells, AKT was found localized in their cytoplasm. TGF-beta did not induce phospho-SMAD3 in these cells. The data indicate that NKX2-1 inhibits SMAD pathway through AKT activation. Ourwork demonstrates that TGF-beta signaling is controlled by NKX2-1 in thyroid stem cells.

Citation Format: Manabu Iwadate, Yoshinori Takizawa, Shioko Kimura. NKX2-1 regulates SMAD and non-SMAD pathways in thyroid stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 357. doi:10.1158/1538-7445.AM2017-357
Abstract 3574: A survey of mutations in biomedical literature using a machine based approach
Authors: Koyama Takahiko , Rhrissorrakrai Kahn , Parida Laxmi
Published: July 2017

Introduction:

Being able to characterize mutations for both pathogenicity and drug response is indispensable to the analysis of tumor genomics and the development of therapeutic options. While a great deal of data has been deposited in various structured, genomic databases, a large portion of insights are primarily and often times solely found in biomedical literature. Medline contains about 26 million literature citations; a number that is unrealistic for a human to read. Thus machine based approaches are needed to comprehensively capture the landscape of reported mutations.

Method:

An automated pattern matching method is utilized to extract mutations from Medline abstracts as presented in Human Genome Variation Society (HGVS) format and RefSNPs (rs) number. A typical HGVS protein mutation is described as [reference amino acid][position][new amino acid], as in p.His1047Arg, His1047Arg, or simply H1047R in HGVS format. This method identifies and consolidates all mentioned protein mutations and their alternate formulations.

Result:

Over 300,000 unique abstract-mutation pairs were identified including 90,000 unique mutations. Well known cancer mutations such as BRAF V600E, JAK2 V617F and EGFR L858R are among the most frequent appearing in oncology literature. At the other end, 51,000 mutations are mentioned in just a single abstract, 16,000 mutations in two abstracts, 7,600 in three abstracts, and so forth.

Conclusion:

The number of mutations appearing in Medline abstracts represents just a small portion of the 2 million unique coding mutations contained in the COSMIC database. While we expect the actual coverage of mutations by literature to be more comprehensive if this approach is extended to the full text body, the number would likely remain small compared with the total reported COSMIC mutations. One of the great challenges in oncology is characterizing variants of unknown significance (VUS), and by first extracting all reported mutations, even those mentioned in only one article, and their specific biological context, we can begin to identify broader patterns in mutations’ pathogenicity and their impact on drug response.

Citation Format: Takahiko Koyama, Kahn Rhrissorrakrai, Laxmi Parida. A survey of mutations in biomedical literature using a machine based approach [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3574. doi:10.1158/1538-7445.AM2017-3574
Abstract 3588: Computational method to identify non-coding cancer drivers
Authors: Liu Eric Minwei , Dhingra Priyanka , Fundichely Alexander M. , Khurana Ekta
Published: July 2017

Cancer is caused by mutations in driver genes - yet ~25% patients do not show any mutations in known drivers. ~99% somatic mutations obtained from whole-genome sequencing localize to regions that do not code for proteins and many of these non-coding mutations regulate gene expression. Although variants in protein-coding regions have received the majority of attention, numerous studies have now noted the importance of non-coding variants in cancer - for example, mutations in the promoter of the TERT gene. Identification of functional non-coding variants that drive tumor growth remains a challenge and a bottleneck for the use of whole-genome sequencing in the clinic. Cancer drivers are generally identified by the high frequency at which their mutations occur across patients. However, mutation rate is highly heterogeneous across the genome and many non-driver elements show higher mutation frequency than others, such as regions bound by transcription factors (TFs) in melanoma or regions replicating late during cell division in colon cancer. We have developed a novel computational approach to predict non-coding cancer drivers that differentiates between regions highly mutated due to positive selection (true drivers) vs those due to mutation rate heterogeneity (false positives). This method integrates the signals of high functional impact of variants with the recurrence of variants across multiple tumor samples to identify the elements that show more and higher functional impact mutations than expected randomly. Our method controls for multiple covariates of mutation rates, such as replication timing, histone modification marks and open chromatin regions to identify the non-coding cancer drivers. The functional impact is predicted using the FunSeq scheme that uses the properties of ENCODE elements (including conservation, TF motif disruption and network properties) within a weighted scoring scheme to predict deleteriousness of non-coding variants. I will present the details of this method and discuss the ongoing efforts to analyze ~2,900 tumor whole-genomes in the ‘Pan-Cancer Analysis of Whole Genomes, PCAWG’ consortium.

Citation Format: Eric Minwei Liu, Priyanka Dhingra, Alexander M. Fundichely, Ekta Khurana. Computational method to identify non-coding cancer drivers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3588. doi:10.1158/1538-7445.AM2017-3588
Abstract 3607: Metastasis-associated protein 1 negatively regulates expression of androgen receptor in ER-positive breast cancer cells
Authors: Kim SeungSu
Published: July 2017

Androgen receptor (AR) is expressed in more than 80% of ER-positive breast cancer and its expression is correlated with better prognosis in ER-positive breast cancer. However, little is known about the regulation of AR expression in breast cancer. We have previously reported that metastasis-associated protein 1 (MTA1), a cancer-promoting gene that is overexpressed in breast cancer, increases the expression of ERα in breast cancer. Here, we investigated the role of MTA1 on AR expression in three different ER-positive breast cancer cell lines. First, small interfering RNA-mediated knockdown of MTA1 increased protein and mRNA expression of AR in MCF7, T47D, and ZR75-1. Overexpression of MTA1 down-regulated expression of AR in these cell lines. Activity of the reporter-encoding AR promoter (-2408 bp ~ +1126 bp) was decreased in the presence of MTA1 in a dose-dependent manner. Analysis of the chromatin immunoprecipitation sequencing data (http://encodeproject.org/ENCODE) revealed that MTA1 may bind to 1 kilo base downstream of AR transcription start site. Together, our results demonstrated that MTA1 negatively regulates expression of AR in breast cancer cells. These results suggest that the negative regulation of AR may be associated with the cancer promoting effects of MTA1, which may provide a therapeutic opportunity against ER-positive breast cancer.

Citation Format: SeungSu Kim. Metastasis-associated protein 1 negatively regulates expression of androgen receptor in ER-positive breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3607. doi:10.1158/1538-7445.AM2017-3607
Abstract 3614: Antibody validation revises estrogen receptor beta research
Authors: Andersson Sandra , Sundberg Mårten , Pristovsek Nusa , Ibrahim Ahmed , Jonsson Philip , Katona Borbala , Clausson Carl-Magnus , Zieba Agata , Ramström Margareta , Söderberg Ola , Williams Cecilia , Asplund Anna
Published: July 2017

The discovery of estrogen receptor β (ERβ/ESR2) in 1996 was a landmark discovery. Its homology with breast cancer pharmacological target and predictive biomarker estrogen receptor α (ERα /ESR1) raised hopes for improved endocrine therapies. However, after twenty years of intense research the therapeutic promises have not materialized. We here perform a rigorous validation of 13 anti-ERβ antibodies, and demonstrate that the vast majority are unspecific. Using well-characterized controls and a panel of validation methods, we conclude that only the rarely used monoclonal antibody PPZ0506 targets ERβ in a specific manner in immunohistochemistry. We applied this antibody for protein expression profiling in 44 normal and 21 malignant human tissues, and found that detection of ERβ protein is limited to testis, ovary, placenta, lymphoid cells, granulosa cell tumors, and a subset of malignant melanoma and thyroid cancers. This expression pattern aligns well with RNA-seq data, but contradicts a multitude of studies. We did not find evidence that normal or cancerous human breast express ERβ protein. Our study highlights how inadequately validated antibodies can lead an exciting and promising field astray.

Citation Format: Sandra Andersson, Mårten Sundberg, Nusa Pristovsek, Ahmed Ibrahim, Philip Jonsson, Borbala Katona, Carl-Magnus Clausson, Agata Zieba, Margareta Ramström, Ola Söderberg, Cecilia Williams, Anna Asplund. Antibody validation revises estrogen receptor beta research [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3614. doi:10.1158/1538-7445.AM2017-3614
Abstract 3619: Role of MTDH in estrogen-regulated gene expression
Authors: Meng Xiangbing , Li Yujun , Bosquet Jesus Gonzalez , yang shujie , Thiel Kristina W. , Leslie Kimberly K.
Published: July 2017

Disruption of estrogen signaling is widely associated with the development of breast, endometrial, and ovarian cancers. As a multifunctional mediator of carcinogenesis, metadherin (MTDH)/AEG-1 overexpression has been associated with numerous types of cancers, with reported roles in tumor initiation, proliferation, invasion, metastasis and chemoresistance. At the molecular level, MTDH has been shown to interact with proteins that drive tumorigenesis, including NF-κB, PLZF, BCCIPα and SND1. Through analysis of the Cancer Genome Atlas (TCGA) datasets for ER-positive endometrial and breast cancers, we found that over 25% of all gene expression correlated with MTDH. Using Affymetrix microarrays, we characterized the differences in gene expression between estrogen-treated parental and MTDH-deficient endometrial and breast cancer cells. We also explored a possible interaction between MTDH and ER by immunoprecipitation and found that MTDH and ER associated in both breast and endometrial cancer cells in response to estrogen. Reciprocal co-immunoprecipitation analysis demonstrated that acute estrogen stimulation promoted the interaction of MTDH with ER in the nucleus. These data suggest that MTDH and ERα interact in the nucleus with estrogen treatment, and MTDH/ER co-regulated genes may have functions in drug resistance, metastasis and tumor progression.

Citation Format: Xiangbing Meng, Yujun Li, Jesus Gonzalez Bosquet, shujie yang, Kristina W. Thiel, Kimberly K. Leslie. Role of MTDH in estrogen-regulated gene expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3619. doi:10.1158/1538-7445.AM2017-3619
Abstract 3627: Validation of PPARG and RXRA as drivers of bladder cancer
Authors: Goldstein Jonathan T. , Strathdee Craig , Duke Fujiko , Shih Juliann , Meyerson Matthew
Published: July 2017

A subset of muscle-invasive bladder cancer (BLCA) is typified by PPARG pathway activation. PPARG focal gene amplification occurs in 15% of bladder cancer patients, and similarly, 5% of BLCA patients possess hotspot mutations in the requisite heterodimer partner of PPARG, RXRA (S427F, S427Y). We used genetic perturbation to study the role of PPARG in bladder cancer. Our results show that overexpression of RXRA and PPARG mutant alleles activate expression of PPARG and PPARA target genes in a ligand-independent manner, and that bladder cancer cell lines are dependent on PPARG for viability. These findings suggest that PPARG may be a promising therapeutic target for treatment of bladder cancer.

Citation Format: Jonathan T. Goldstein, Craig Strathdee, Fujiko Duke, Juliann Shih, Matthew Meyerson. Validation of PPARG and RXRA as drivers of bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3627. doi:10.1158/1538-7445.AM2017-3627
Abstract 365: Cross-talk between BRAF and Hippo/YAP1 signaling in melanoma
Authors: Raj Mohan Kumar Durai , Nguyen Jonathan , Bora-singhal Namrata , Messina Jane , Gibney Geoffrey , Chellappan Srikumar
Published: July 2017

Hippo/YAP1 signaling pathway is a tumor suppressive pathway that controls the organ size by modulating the cell growth, proliferation and apoptosis and is conserved from Drosophila to mammals. In mammals, the Hippo tumor suppressor pathway consists of cascade of kinases in which MST1/2 phosphorylates and activates LATS1/2. The latter phosphorylates the oncogenic transcriptional coactivators YAP1 and TAZ, leading to their cytoplasmic retention by 14-3-3 proteins and/or degradation. Inactivation of MST and LATS kinases allows YAP1 and/ or TAZ nuclear translocation and subsequent activation of their target genes.

Deregulation of Hippo pathway can induce tumors in model organisms and occurs in wide range of human cancers including melanoma. Merlin, a key component of this pathway which inhibits YAP1, is mutated/deleted in 8% of melanoma. Majority of uveal melanomas are driven by Gq/11 mutations that trigger YAP1 nuclear translocation, promoting tumor growth. High levels of YAP1 in BRAFV600E mutant tumors confer resistance to RAF- and MEK- targeted therapy in patients.

Our results show that YAP1 level is elevated in melanoma and the YAP1 inhibitor verteporfin alone or in combination with B-RAF inhibitor PLX4720 reduces the viability, invasion and anchorage-independent growth of B-RAF V600E mutant SK-MEL-28 and SK-MEL-5 cells. In addition, verteporfin treatment also reduced the viability of PLX4720 resistant 1205 cells. Western blot analysis of verteporfin and PLX4720 treated SK-MEL-28 and SK-MEL-5 cells displayed reduced levels of YAP1, B-RAF, pERK, MEK and pMEK.

We also report a novel physical interaction between YAP1 and B-RAF; this could be detected using double immunofluorescence and immunoprecipitation-western blotting techniques in both B-RAF V600E mutant and N-Ras mutant melanoma cells. Proximity ligation assays on tissue microarray showed that YAP1-B-RAF interaction is elevated in metastatic melanoma compared to normal skin. These novel findings highlight the crosstalk between B-RAF and Hippo/YAP1 signaling which might have a

potential role in melanoma development and progression. Further, experiments are in progress to elucidate the functional significance of YAP1-B-RAF interaction in melanoma.

Citation Format: Mohan Kumar Durai Raj, Jonathan Nguyen, Namrata Bora-singhal, Jane Messina, Geoffrey Gibney, Srikumar Chellappan. Cross-talk between BRAF and Hippo/YAP1 signaling in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 365. doi:10.1158/1538-7445.AM2017-365
Abstract 3668: Oncolytic adenoviruses expressing OX40L or GITRL immune modulators show antitumor effect on immune-competent mouse breast cancer models
Authors: Martinez Francisco W. Puerta , Rivera Yisel A. , Nguyen Teresa , Fan Xuejun , Henderson Jared M. , Rehnuma Shifat , Hossain Mohammad B. , Jiang Hong , Fueyo Juan , Manzano Candelaria Gomez
Published: July 2017

Metastasis of advanced stage cancers remains as the main cause of morbidity and mortality in oncologic patients. Metastatic cancers, especially those that metastasize to the brain, are generally resistant to conventional therapies. Thus, more innovative and efficacious therapies are urgently needed. Therapeutic goals are the specific targeting of malignant cells, shrinkage of stablished tumors, prevention and/or eradication of metastases and, ultimately, induction of a specific anti-tumor immune response. In this study we tested the efficiency of a treatment regimen consisting of oncolytic adenoviruses combined with specific immune regulators to prevent tumor progression and metastasis. In order to do so, mouse metastatic breast cancer cells 4T1 or 66c14, were orthotopically implanted in female BALB/c mice. The resulting primary tumors were treated with multiple doses of third generation adenoviruses targeting different immune checkpoints, such as, OX40/OX40L and GITR/GITRL pathways in the immune synapse. The treatment with these adenoviral constructs resulted in T cells activation and reduction of the metastases in 50% of the mice. In addition, the size and number of the metastases detected in lungs were significantly lower comparing with those observed in the control groups. Survival rates were also significantly different (P < 0.046) when compared to controls. FACS analysis of isolated tumor-infiltrating lymphocytes, as well as, splenocytes showed that OX40/OX40L or GITR/GITRL immune therapies were able to activate and increase the number of cytotoxic T cells and suppress tumor anergy. In summary, we demonstrated for the first time that Delta-24-RGDOX and Delta-24-GREAT, expressing OX40L and GITRL respectively, display antitumor effect on immune competent models of metastatic breast cancer.

Citation Format: Francisco W. Puerta Martinez, Yisel A. Rivera, Teresa Nguyen, Xuejun Fan, Jared M. Henderson, Shifat Rehnuma, Mohammad B. Hossain, Hong Jiang, Juan Fueyo, Candelaria Gomez Manzano. Oncolytic adenoviruses expressing OX40L or GITRL immune modulators show antitumor effect on immune-competent mouse breast cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3668. doi:10.1158/1538-7445.AM2017-3668
Abstract 3711: R-Ras is required for T cell trafficking in high endothelial venues and implicated in anti-tumor immunity
Authors: Chan Andrew M. , Yan Xiaocai , Liu Wai Nam , Yan Mingfei
Published: July 2017

R-Ras belongs the RAS superfamily of small GTP-binding proteins implicated in cell adhesion signaling. Unlike classical Ras proteins, R-Ras is not activated by growth factor receptors. Rather, its GTPase activity is modulated by molecules implicated in cell-cell adhesion including semaphorins and notch. Using an R-Ras knockout mouse strain, we have previously reported that R-Ras is required for murine Dendritic cell functions. Further characterization of this mouse strain revealed reduced cellularity of peripheral lymph nodes (PLNs) by 40% with T cell regions in the paracortex in which high endothelial venules (HEV) resided were less prominent when compared with wild-type (WT) mice. Consistently, immunohistological analysis for MECA-79, a marker of mature HEV, showed a 3- to 4-fold decrease in staining intensity. This is correlated with a 30-40% reduction in the number of CD62L+ CD4 and CD8 T cells in PLNs. Furthermore, R-Ras null T cells had reduced proliferative and homing capacity when compared with WT. More importantly, we demonstrated for the first time that the chemokine CCL21 was able to stimulate the GTP-loading of R-Ras within 15 seconds of addition. Indeed, R-Ras null T-cells displayed reduced binding to soluble ICAM-1 as well as to 2H-11 endothelial cells. To explore the role of R-Ras in anti-tumor immunity, B16/F10 melanoma cells were injected subcutaneously to WT and R-Ras null mice. In addition, a mouse breast tumor model was generated ineither WT or R-Ras-null genetic background. Tumor development and the number of infiltrating immune cells were analyzed to examine if R-Ras was involved in either anti- or pro-tumor immune responses. This work was supported by NIH CA78509 (AMC), MH59771 (AMC). MY was supported by Hong Kong PhD Fellowship Scheme (PF12-13584) and AMC was supported by the Lo Kwee-Seong Seed Fund, and a General Research Fund Grant, 14120915, from Hong Kong University Grants Committee.

Note: This abstract was not presented at the meeting.

Citation Format: Andrew M. Chan, Xiaocai Yan, Wai Nam Liu, Mingfei Yan. R-Ras is required for T cell trafficking in high endothelial venues and implicated in anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3711. doi:10.1158/1538-7445.AM2017-3711
Abstract 3719: Bicyclic peptides for PET imaging of MT1-MMP expressing tumors
Authors: Teufel Daniel , Harrison Helen , Campbell Spencer , Stace Catherine , Walker Edward , Lutz Robert J. , Park Peter , Eder Matthias , Bauder-Wüst Ulrike , Schierbaum Ursula , Leotta Karin , Kopka Klaus , Haberkorn Uwe
Published: July 2017

The Bicycle® technology is based on repertoires of short peptides displayed on the surface of bacteriophages which can be modified with homo-trifunctional organochemical scaffolds, thus creating large diverse libraries of constrained, bicyclic peptides. These large combinatorial libraries have been extensively used for iterative selections to identify high affinity binding peptides to a wide array of targets, including receptors, interleukins and proteases.

Bicyclic peptides are chemically synthesized macrocyclic entities with drug-like properties that exhibit sub-nanomolar affinities and exquisite selectivity towards targets. Unlike biologics, their synthetic nature allows facile modulation of metabolic and pharmacokinetic properties, as well as site-specific conjugation to effector molecules such as fluorophores, radionuclides, and cytotoxic drugs.

In the present work, novel phage display derived bicyclic peptides were identified targeting the matrix metalloproteinase 14 (also known as MT1-MMP), a tumor associated surface protein overexpressed in a variety of cancers (i.e. lung, breast). A prototype bicyclic peptide with high affinity to MT1-MMP (Kd at ~1 nM) was identified, and confocal microscopy using fluoresceinated bicyclic peptide derivatives shows target-dependent internalisation in MT1-MMP+ cells. In the in vivo mouse, selective tumor binding in an MT1-MMP+ xenograft model is demonstrated for a DOTA conjugate loaded with Ga-68 or Lu-177. Upon proteolytic optimization of the prototype bicycle peptide, a striking enhancement in tumor signal is observed in biodistribution studies. Compared to radiolabeled antibodies directed against the same target, the lead compound showed fast background clearance (< 1 %ID/g for all organs apart from kidneys) resulting in high imaging contrast in µPET studies as early as 30 minutes post injection. Importantly, most of the non-tumor associated bicyclic peptide rapidly clears into the bladder.

Together, tumor targeting bicyclic peptides can, through their small size and high selectivity, facilitate efficient penetration and visualization of tumors in vivo, demonstrating their potential as diagnostic imaging agents in profiling and therapeutic management of patients.

Citation Format: Daniel Teufel, Helen Harrison, Spencer Campbell, Catherine Stace, Edward Walker, Robert J. Lutz, Peter Park, Matthias Eder, Ulrike Bauder-Wüst, Ursula Schierbaum, Karin Leotta, Klaus Kopka, Uwe Haberkorn. Bicyclic peptides for PET imaging of MT1-MMP expressing tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3719. doi:10.1158/1538-7445.AM2017-3719
Abstract 3728: Imaging biomarkers of aromatase-inhibitor induced joint pain
Authors: Goldstein Brian A. , Taljanovic Mihra , Chalasani Pavani , Kwoh C. Kent , Hadden Amanda , Witte Russell , Martinez Jessica
Published: July 2017

Estrogen receptor positive (ER+) breast cancer patients undergoing treatment with aromatase inhibitors (AI) tend to discontinue the supplement when to painful side effects appear. It is believed that women with AI-induced musculoskeletal syndrome (AIMSS) have specific physiological conditions that differentiate them from women with normal aging-related inflammatory processes (e.g., rheumatoid arthritis or RA). Ultrasound shear wave elastography (USWE) provides a real-time, quantitative assessment of the elastic modulus of soft tissue, which is a potential biomarker that is altered in women with AIMSS.

Because USWE is a completely noninvasive technique available on state-of-the-art clinical ultrasound scanners, it can be readily applied to study patients who undergo AI treatment and help predict adverse effects during therapy.

A Siemens s3000 clinical scanner and 9 MHz linear probe were used for USWE of the hands and wrists in postmenopausal women on AI, postmenopausal women with RA and healthy postmenopausal women. The scans included an evaluation of major wrist tendons, nerve, and joints with shear wave velocities ranging from 0.5 to 15 m/s, which are directly related to the local shear modulus – an absolute measure of tissue stiffness. The ultrasound images and raw shear wave data were then analyzed in MATLAB-TM using a custom graphical user interface (GUI) to calculate the distribution of velocities (or shear moduli) in a user-defined region of interest (ROI). The results were exported to a spreadsheet in Excel for further analysis and comparison among the different groups.

Initial results suggest the shear wave velocity and estimated Young’s modulus (λ) were significantly higher in AI patients (14.6 ± 0.2 m/s, λ=642 ± 17 kPa) compared to RA (6.84 ± 1.1 m/s, λ=143 ± 43 kPa) and healthy (7.58 ± 0.9 m/s, λ=174 ± 42 kPa) patients. These values suggest that the mechanical properties of tendons in the wrist may change during treatment with AI and contribute to joint pain in these subjects. It is critical that ER+ patients on AI complete their full treatment. As a potential biomarker of AIMSS and associated pain, USWE may help predict which patients are most susceptible to these side effects, promote early intervention to reduce or eliminate symptoms, and help increase adherence to AI therapy.

Citation Format: Brian A. Goldstein, Mihra Taljanovic, Pavani Chalasani, C. Kent Kwoh, Amanda Hadden, Russell Witte, Jessica Martinez. Imaging biomarkers of aromatase-inhibitor induced joint pain [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3728. doi:10.1158/1538-7445.AM2017-3728
Abstract 373: Wnt signaling induces anti-apoptotic effect in colorectal cancer cells through the suppression of IFITs
Authors: Ohsugi Tomoyuki , Yamaguchi Kiyoshi , Zhu Chi , Ikenoue Tsuneo , Furukawa Yoichi
Published: July 2017

Impaired Wnt signaling pathway plays a crucial role in the development of colorectal cancer through the activation of β-catenin/TCF complex. Although genes up-regulated by Wnt/β-catenin signaling has been well studied, the down-regulated genes are poorly understood. To clarify the comprehensive changes regulated by the signaling in colorectal cancer cells, we explored a global gene expression of CRC cells transfected with β-catenin siRNAs or dominant negative form of TCF7L2 (dnTCF7L2). Consequently, a set of genes that were negatively regulated by β-catenin/TCF were identified. Among the genes, three members of interferon-induced proteins with tetratricopeptide repeats (IFIT) family (IFIT1, IFIT2, and IFIT3) expression were significantly increased by the inhibition of β-catenin/TCF. Comparison of gene expression data from normal colonic mucosa and the tumor tissues showed that the expression of IFIT1 and IFIT2 in the tumors was significantly lower than that in normal tissues. To elucidate the mechanism of IFITs expression regulated by β-catenin/TCF, we performed a reporter assay using plasmid containing 1.2-kb of 5’-flanking region of the IFIT2 gene. As a result, the reporter activity was significantly enhanced by either transduction of β-catenin or dnTCF7L2, suggesting that blockage of β-catenin/TCF stimulated IFITs through the promoter. In addition, we found that overexpression of IFIT2 increased apoptosis and decreased cell proliferation in SW480 and HCT116 cells. These results imply that Wnt signaling may promote anti-apoptotic effect in cancer cells through the suppression of IFIT2. Our findings suggest that analysis of down-regulated genes in response to activated Wnt/β-catenin signaling provides a better understanding of human colorectal carcinogenesis.

Citation Format: Tomoyuki Ohsugi, Kiyoshi Yamaguchi, Chi Zhu, Tsuneo Ikenoue, Yoichi Furukawa. Wnt signaling induces anti-apoptotic effect in colorectal cancer cells through the suppression of IFITs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 373. doi:10.1158/1538-7445.AM2017-373
Abstract 3743: Magnetic resonance imaging shrinkage patterns after neoadjuvant chemotherapy for breast carcinomas: Correlation with molecular subtypes
Authors: Mori Tsuyoshi , Tomida Kaori , Kitamura Mina , Sakai Sachiko , Kawai Yuki , Shimizu Tomoharu , Umeda Tomoko , Ueki Tomoyuki , Kaida Sachiko , Miyake Toru , Iida Hiroya , Akabori Hiroya , Kitamura Naomi , Yamaguchi Tsuyoshi , Sonoda Hiromichi , Naka Shigeyuki , Tani Masaji
Published: July 2017

Background: Neoadjuvant chemotherapy (NAC) is widely performed for patients with locally advanced breast carcinomas. It is important to obtain precise information using imaging modalities about the distribution of residual carcinomas after NAC to predict the success of breast conserving surgery. However, the relationship between magnetic resonance imaging (MRI) shrinkage patterns and molecular subtypes of breast carcinomas has not yet been investigated.

Methods: We analyzed the MRI shrinkage and pathological patterns of residual carcinomas after NAC in 54 consecutive breast carcinomas. The shrinkage patterns were classified into five categories: Type I and II (concentric shrinkage with or without any surrounding lesion), type III (shrinkage with residual multinodular lesions), type IV (diffuse contrast enhancement in the entire quadrant), and non-visualization.

Results: The most common MRI shrinkage pattern was type I (39%) followed by non-visualization (26%). The most common pathological shrinkage pattern was type III (33%), followed by type II (28%) and non-visualization (22%). The concordance rates of the MRI patterns of non-visualization and type I were significantly lower than those of the MRI patterns of type II, III, and IV (p = 0.018). Moreover, in the luminal subtypes, the concordance rates of the MRI patterns of non-visualization and type I were significantly lower than those of the MRI patterns of type II, III, and IV (p = 0.026); however, in the non-luminal subtypes, this correlation was not significant. Conclusions: The results of the present study suggest that the cases with MRI shrinkage patterns of non-visualization and type I require caution regarding the surgical margins compared with the other types, especially the luminal subtypes.

Citation Format: Tsuyoshi Mori, Kaori Tomida, Mina Kitamura, Sachiko Sakai, Yuki Kawai, Tomoharu Shimizu, Tomoko Umeda, Tomoyuki Ueki, Sachiko Kaida, Toru Miyake, Hiroya Iida, Hiroya Akabori, Naomi Kitamura, Tsuyoshi Yamaguchi, Hiromichi Sonoda, Shigeyuki Naka, Masaji Tani. Magnetic resonance imaging shrinkage patterns after neoadjuvant chemotherapy for breast carcinomas: Correlation with molecular subtypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3743. doi:10.1158/1538-7445.AM2017-3743
Abstract 376: Biospecimen and data resources for cancer research from NCI’s BPV program
Authors: Guan Ping , Moore Helen M.
Published: July 2017

The Biospecimen Preanalytical Variables (BPV) Program was initiated by the National Cancer Institute’s Biorepositories and Biospecimen Research Branch to evaluate the impact of preanalytical factors on the molecular integrity of biospecimens. Selected preanalytical factors including cold ischemic time (delay to formalin fixation (DTF)), time in formalin (TIF), freezing methods, and storage temperatures and durations were examined for their potential effects on molecular profiles from surgical resection tissues and matched blood from four cancer types (kidney, ovary, colon and lung). The BPV program has collected tumor specimens from 364 cancer patients. Each specimen was annotated with 300+ data elements that cover steps in the collection, handling, and processing procedures, pathology review, and clinical information. NCI conducted multiple studies using these specimens to evaluate the preanalytical impacts on different analytical platforms including gene expression profiling, copy number variation, proteomics and metabolomics profiling. The program invites interested organizations to work with NCI through collaboration to further evaluate preanalytical effects on molecular analyses (https://techtransfer.cancer.gov/availabletechnologies/e-000-2013). The remaining specimens are available to support relevant research focusing on biospecimen science and/or clinical biomarker assay development (https://specimens.cancer.gov/search/). The IT infrastructure that was developed to support BPV biospecimen collection and management has been further developed into open source products (https://github.com/NCIP/CDR and https://github.com/NCIP/CDR-Lite). The controlled vocabulary that records the terms and definitions used in describing the overall biospecimen collection efforts has been refined and published at publicly accessible CDE repositories: NCI’s caDSR (https://cdebrowser.nci.nih.gov/CDEBrowser/ ) and NIH’s CDE portal (https://cde.nlm.nih.gov/cde/search ). An ongoing collaboration with an academic ontology consortium will map the CDEs to existing biobanking ontology frameworks and make them publicly available. The BPV program has generated a wide range of ~omics data. We are preparing a BPV data compendium to be submitted to dbGaP at NCBI. These data will be used as the experimental evidence to develop evidence-based best practices for fit-for-purpose collection, processing, and storage of biospecimens for cancer research.

Citation Format: Ping Guan, Helen M. Moore. Biospecimen and data resources for cancer research from NCI’s BPV program [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 376. doi:10.1158/1538-7445.AM2017-376