bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract CN07-02: Epigenetics: A gatekeeper to DNA amplification
Authors: Whetstine Johnathan R.
Published: August 2018

DNA copy gains and amplifications are often associated with tumorigenesis, poor prognosis, and drug resistance. However, little is known about how these regions of the genome are selected for amplification. Our group discovered that modulation of epigenetic factors and their associated chromatin states controls DNA copy gains of drug-resistant regions. In fact, transient site-specific copy number gains occur in both normal and cancer cells upon manipulation of a histone H3 lysine 9/36 (H3K9/36) tri-demethylase KDM4A. However, the mechanism by which KDM4A is targeted to specific genomic regions and whether additional chromatin regulators are involved in this process were unknown. To begin addressing these questions, we performed a series of unbiased genetic screens against all the histone lysine-modifying enzymes and followed their ability to generate copy gains of regions using DNA fluorescence in situ hybridization. Our screen uncovered insights about how KDM4A targeting influences DNA amplification in the human genome. We also demonstrated that specific epigenetic factors are working as a network to modulate DNA amplification of drug-resistant regions of the genome. Furthermore, we uncovered KDM4A-independent DNA amplifications, which established that additional regions are undergoing epigenetic control for amplification. These data will be discussed at the meeting.

Citation Format: Johnathan R. Whetstine. Epigenetics: A gatekeeper to DNA amplification [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr CN07-02.
Abstract KN02: Keynote Lecture: T-cell recognition of human cancer
Authors: Schumacher Ton
Published: August 2018

Many human tumors contain large numbers of mutations, of which many hundreds can be present within expressed genes. As the resulting altered protein sequences are foreign to the immune system, immune recognition of such "neoantigens" has been proposed as a major factor in the activity of clinically used immunotherapeutics such as anti-CTLA-4 and anti-PD-1.

In support of this hypothesis, work by others and us over the past years has demonstrated that CD4 and CD8 T-cell recognition of the consequences of DNA damage is a common feature in, e.g., human melanoma, and can also be boosted by immunotherapy.

These data establish "tumor foreignness," as determined by neoantigen formation, as one of the determinants of effective cancer immunotherapies. However, it is evident that the outcome of cancer-immune interaction is also critically dependent on a number of other factors. With the ultimate aim to describe a "cancer immunogram" that depicts the state of cancer-immune interactions for individual patients, we are developing and exploiting genetic screens and (functional) assays of human tumor material. A detailed understanding of the specific bottleneck(s) in immune-mediated tumor control in individual patients should be of value for both patient selection and for the development of novel clinical interventions.

Citation Format: Ton Schumacher. Keynote Lecture: T-cell recognition of human cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr KN02.
Abstract LB-A15: An Ex-vivo Platform Predicts Anti-tumor Outcome of Metabolically-Targeted, Algorithm-Driven Combination Therapy in Triple-Negative Breast Cancer
Authors: Goldman Aaron J.
Published: August 2018

Abstract: Cancer cells undergo phenotypic cell state transitions in response to chemotherapy as a mechanism that can confer transient resistance. However, such cell state transitions can also unlock unique vulnerabilities that can be exploited using temporally-sequenced combination chemotherapy. Here, utilizing a primary breast cancer ex-vivo functional assay (CANScriptTM) that captures tumor heterogeneity, we report that in response to a chemotherapeutic agent, a subset of cancer cells can mount an acutely-induced phenotypic adaptive resistance to future cytotoxic pressure via the transient acquisition of a unique metabolic state defined by augmented glycolysis together with mitochondrial proficiency. These cells activate two complex, temporally-interdependent pathways that enable a glucose shunt towards the pentose phosphate pathway (PPP), which confers an adaptive cross-tolerance to different chemotherapeutic agents. Mathematically modeling these pathways, and simulating drug schedules, we define a rationally-designed 3-drug combination therapy of metabolic inhibitors and cytotoxic agents, which results in improved cancer survival. Our findings highlight a new bioenergetics-based adaptive resistance mechanism through which cancer cells can survive combinations of chemotherapy. Administration of metabolic inhibitors in rational, temporal sequence with existing chemotherapy can emerge as a new paradigm in the treatment of cancer.

Citation Format: Aaron J. Goldman. An Ex-vivo Platform Predicts Anti-tumor Outcome of Metabolically-Targeted, Algorithm-Driven Combination Therapy in Triple-Negative Breast Cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A15.
Abstract LB-A16: Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer
Authors: Dijkmans Thomas
Published: August 2018

Creating patient-derived xenograft (PDX) models from human cancers allows bio-banking, propagation, characterization and drug testing on a large panel of tumor variants. To scale up the number of compound tests, identify relevant compound doses, increase the throughput, and study relevant cancer-driving pathways we developed 3D in vitro culture methods for PDX model-derived cell isolates. In this effort, each individual model is subjected to a wide panel of culture medium formulations, amongst which some media contain WNT-pathway stimulating components. In one of our campaigns we identified an absolute dependency of one of the tested Pancreatic in vitro PDX models to the WNT-signaling potentiating factor R-Spondin, while other Pancreas models showed little or no dependency. To further study the dependency of Pancreatic PDX models on the functionality and signaling of the WNT-pathway, we exposed these in vitro PDX models to increasing doses of bio-similars of 18R5 (anti-FZD7), 54F28 (FZD8-Fc decoy receptor) and to the Porcupine inhibitor LGK974, which blocks WNT palmitoylation and therefore secretion. Our findings are that the WNT-pathway is functional and contributing to Pancreas cancer outgrowth and morphological differentiation at different extents and levels in different patient-models. Our in vitro PDX culture system in combination with 3D phenotypic analysis allows in-detail study of the WNT-pathway, as well as by supplementation or omission of secreted factors, addition of small molecule inhibitors and by exposure to antibodies. These in vitro PDX models can be used to test novel compounds and antibodies targeting the WNT pathway.

Citation Format: Thomas Dijkmans. Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A16.
Abstract LB-A23: Discovery of a potent catalytic p300/CBP inhibitor that targets lineage-specific tumors
Authors: Bromberg Kenneth D. , Lasko Loren M. , Jakob Clarissa G. , Qiu Wei , Montgomery Debra , Digiammarino Enrico L. , Hansen Todd M. , Risi Roberto M. , Frey Robin R. , Manaves Vlasios , Shaw Bailin , Algire Mikkel , Hessler Paul , Lam Lloyd T. , Uziel Tamar , Favire Emily , Ferguson Debra , Buchanan Fritz G. , Martin Ruth L. , Torrent Maricel , Rosenberg Saul H. , Michaelides Michael R. , Lai Albert
Published: August 2018

The dynamic and reversible acetylation of proteins catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs) is a major epigenetic regulatory mechanism of gene transcription associated with multiple diseases. While HDAC inhibitors are approved to treat certain cancers, progress on the development of drug-like HAT inhibitors has lagged. The HAT paralogs p300 and CBP (p300/CBP) are key transcriptional co-activators essential for a multitude of cellular processes and also implicated in human pathological conditions, including cancer. Current p300/CBP HAT domain inhibitors including natural products, bi-substrate analogs (Lys-CoA) and the widely utilized C646 lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like p300/CBP catalytic inhibitor. We show the first high resolution (1.95Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 is acetyl-CoA competitive. A-485 selectively inhibited proliferation across lineage-specific tumor types, including several hematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen sensitive and castrate resistant prostate cancer and inhibited tumor growth in a castration resistant xenograft model. These results demonstrate the feasibility of selectively targeting the catalytic activity of histone acetyltransferases.

Citation Format: Kenneth D. Bromberg, Loren M. Lasko, Clarissa G. Jakob, Wei Qiu, Debra Montgomery, Enrico L. Digiammarino, Todd M. Hansen, Roberto M. Risi, Robin R. Frey, Vlasios Manaves, Bailin Shaw, Mikkel Algire, Paul Hessler, Lloyd T. Lam, Tamar Uziel, Emily Favire, Debra Ferguson, Fritz G. Buchanan, Ruth L. Martin, Maricel Torrent, Saul H. Rosenberg, Michael R. Michaelides, Albert Lai. Discovery of a potent catalytic p300/CBP inhibitor that targets lineage-specific tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A23.
Abstract LB-A26: Regulation of super-enhancers by GZ17-6.02 has therapeutic effect in pancreatic cancer (PDAC)
Authors: Ghosh Chandrayee , Gunewardena Sumedha , Paul Santanu , West Cameron , Dhar Animesh
Published: August 2018

Super enhancers (SE) are defined as unique areas of the genome that are densely bound by numerous transcription factors and play a pivotal role in the cell, including, tissue specification, identity and maintenance. Super enhancers are known to regulate the expression of associated genes and often drive high-level transcription. Studies have established many genes that play an important role in cancer biology as SE-driven oncogenes. It is likely that the peculiar pancreatic-specific tumor phenotype is a consequence of oncogenes hacking the resident tissue regenerative program, thus interfering with super enhancer-driven repair networks exerting a disproportionately disruptive effect on tumor versus normal pancreatic tissue. In this study, we have used ChIP-seq, RNA-seq and bioinformatics analysis to identify super-enhancers in the human metastatic pancreatic cancer cell line S2-007. H3K27ac marks were identified at enhancer region of numerous genes that act as SE in PDAC. The most prominent super-enhancers identified, based on a high level of H3K27ac marks were associated with c-MYC, MED1, OCT-4, NANOG and SOX2 genes. Previously we showed that “isovanillin”, one of the major compound of A. palaestinum, in combination with other anticancer agents termed as ‘GZ17-6.02’ inhibited tumor progression and metastasis in PDAC by suppressing cancer stem cells (CSCs) and sonic hedgehog (SHH) pathway in both in vitro and in vivo systems. In this study, we found that GZ17-6.02 affects acetylation of histones at some of the major SE related genes and at a high dose, a complete reduction in acetylation marks was seen in embryonic stem cell transcription factors. The mRNA sequencing data after GZ17-6.02 treatment also shows a reduction in transcription of, major transcription factors, SHH pathway genes, and stem cell markers. Cancer cells are more sensitive to lower doses of GZ17-6.02 than normal cells; hence normal cells are expected to have fewer side effects. There is no systematic study showing an association of super-enhancers with pancreatic cancer so far. Hence, in the present study, we have established that several super-enhancer targets can be targeted by GZ17-6.02 in PDAC. This study concludes that super-enhancers can be an important therapeutic target for PDAC.

Citation Format: Chandrayee Ghosh, Sumedha Gunewardena, Santanu Paul, Cameron West, Animesh Dhar. Regulation of super-enhancers by GZ17-6.02 has therapeutic effect in pancreatic cancer (PDAC) [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A26.
Abstract LB-B03: Exosome-based Detection of EGFR T790M in Plasma from Non-Small Cell Lung Cancer Patients
Authors: Castellanos Elena , Grimm Dominik G. , Tadigotla Vasisht , Bentink Stefan , Hurley James , Healy John , Neal Patricia L. , Sher Mia , Baughman Dan , Karlovich Chris , Raponi Mitch , Krug Anne , Noerholm Mikkel , Raez Luis E. , Skog Johan
Published: August 2018

About 60% of non-small cell lung cancer (NSCLC) patients develop resistance to targeted epidermal growth factor receptor (EGFR) inhibitor therapy through the EGFR T790M mutation. Although these patients respond well to third generation tyrosine kinase inhibitors like osimertinib, obtaining tissue biopsies for mutation profiling poses risks and is often not feasible. Detecting the mutation in a liquid biopsy from plasma has emerged as a viable alternative to biopsy, however existing liquid biopsy tests using cell-free DNA (cfDNA) often suffer from low sensitivity due to the low copy number of mutant molecules in circulation. As an example, the Food and Drug Adminstration (FDA) approved companion diagnostic for T790M mutation status in plasma (cobas® EGFR mutation Test version 2) only has a 58% sensitivity at a specificity of 80%. In this study we aimed to achieve a higher performance by combining the mutated alleles on cfDNA as well as exosomal RNA and DNA (exoNA) .

ExoNA and cfDNA were extracted from the plasma of NSCLC patients with biopsy-confirmed T790M-positive (N = 102) or T790M-negative (N = 108) samples. The T790M mutation status in plasma exoNA and cfDNA was determined using an analytically validated allele-specific qPCR assay in a CLIA laboratory.

The training cohort consisted of 105 samples (51 positives, 54 negatives). The validation cohort consisted of 105 samples (51 positives, 54 negatives), where ~40% of the patients had intrathoracic disease(M0/M1a). Detection of the T790M mutation in exoNA and cfDNA achieved 92% sensitivity and 89% specificity when compared to tumor biopsy results in the validation cohort. We also observed a very high sensitivity (88%) in patients with intrathoracic disease (M0/M1a), patients for whom detection by liquid biopsy has been proven particularly challenging.

The combination of exoNA and cfDNA for EGFR T790M detection has a higher sensitivity and specificity compared to historical cohorts using ctDNA alone. This is especially important as it could aid giving the right treatment to patients where biopsy is not available or avoid unnecessary tumor biopsies for T790M mutation testing.

Citation Format: Elena Castellanos, Dominik G. Grimm, Vasisht Tadigotla, Stefan Bentink, James Hurley, John Healy, Patricia L. Neal, Mia Sher, Dan Baughman, Chris Karlovich, Mitch Raponi, Anne Krug, Mikkel Noerholm, Luis E. Raez, Johan Skog. Exosome-based Detection of EGFR T790M in Plasma from Non-Small Cell Lung Cancer Patients [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B03.
Abstract LB-B05: Targeting IDH1 inhibits survival and growth of pancreatic cancer cells
Authors: Vaziri-Gohar Ali , Dukleska Katerina , Brody Jonathan R. , Winter Jordan M.
Published: August 2018

Pancreatic ductal adenocarcinoma (PDA) cells have elevated levels of reactive oxygen species (ROS) compared to adjacent normal cells. Although ROS play a role in pancreatic carcinogenesis, the accumulation of excess ROS at later stages of cancer development threaten PDA cell viability. Therefore, adaptive mechanisms to neutralize excess ROS are critical for PDA cell survival. Recently, we discovered the regulatory RNA binding protein, HuR (ELAVL1), protects PDA cells under oxidative stressors, like chemotherapy and nutrient deprivation, by upregulating the NADPH-generating enzyme, isocitrate dehydrogenase 1 (IDH1) through post-transcriptional stabilization of the RNA transcript. This action enhances PDA reductive power under oxidative stress. We hypothesized that IDH1 would be an effective target to treat PDA under its native harsh microenvironment. We deleted IDH1 from isogenic PDA cells using CRISPR gene editing technology and studied the phenotypic impact under physiologic stress. Under both normal culture conditions and nutrient deprivation, IDH1-/- cells had increased ROS as measured by a DCFDA assay, and decreased cell viability as measured by the PicoGreen DNA quantitation assay. Furthermore, IDH1-/- cells were more susceptible to chemotherapy treatment in vitro (gemcitabine or 5-FU) compared with control cells. In vivo experiments revealed that IDH1-/- cells have reduced ability to engraft and grow as xenografts. Taken together, these results suggest that targeting wild type IDH1 is a compelling therapeutic strategy to treat PDA cells, particularly in the context of their extreme and austere microenvironment.

Citation Format: Ali Vaziri-Gohar, Katerina Dukleska, Jonathan R. Brody, Jordan M. Winter. Targeting IDH1 inhibits survival and growth of pancreatic cancer cells [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B05.
Abstract LB-B06: p65BTK is a novel theranostic target in NSCLC
Authors: Grassilli Emanuela
Published: August 2018

Recently we identified new targets to overcome drug resistance in colon cancer; in particular, we discovered and characterized p65BTK, a novel isoform of the Bruton Tyrosine Kinase (BTK). p65BTK is a novel oncogene and a pivotal downstream effector of RAS; its inhibition affects growth and survival of colon cancer cells, (Grassilli et al, Oncogene 2016) and restore the response to chemotherapy in a p53-null background. Here, we investigated whether p65BTK might be a therapeutic target also in Non Small Cell Lung Cancer (NSCLC). Immunohistochemistry assays using anti-p65BTK specific polyclonal antibodies raised in the lab were performed to study p65BTK expression in NSCLC patient specimens. In order to determine the effect of p65BTK inhibition on cell biology, proliferation and viability of NSCLC cell lines with different genetic backgrounds, we carried out growth curves and colony formation assay in presence of increasing concentrations of different BTK inhibitors (Ibrutinib, AVL-292, RN-486). In order to determine whether p65BTK could be a novel theranostic target in NSCLC, representative cell lines were treated with chemotherapy (Cisplatin, Gemcitabine, Pemetrexed) or EGFR-targeted therapy (Gefitinib, Erlotinib) alone or in combination with non-toxic concentrations of BTK inhibitors and then their viability was assessed. First, we showed that p65BTK is expressed in 50% of NSCLC patients’ tissue and its expression correlates with histotype, smoke habit and wtEGFR. In fact, p65BTK is significantly more expressed in adenocarcinoma vs squamous carcinoma patients and in non-smoker vs smoker patients. Finally, its expression is significantly higher in non-smoker patients bearing wild type vs mutant EGFR. Then, we demonstrated that NSCLC cell lines scarcely sensitive to EGFR-targeted therapy overexpress active p65BTK and their viability of is affected in a dose-dependent manner by BTK inhibition. In fact, p65BTK inhibition strongly impairs proliferation and clonogenicity of the same NSCLC cell lines. Notably, p65BTK inhibition reverts resistance of NSCLC cell lines to Cisplatin and Gefitinib, independently of the EGFR and K-Ras status. In conclusion, our data indicate that the inhibition of p65BTK resensitizes NSCLC cells scarcely responsive to chemotherapy and targeted therapy, suggesting that the combined treatment of BTK inhibitors and classic chemotherapy or targeted therapy could represent a novel approach to overcome drug resistance.

Citation Format: Emanuela Grassilli. p65BTK is a novel theranostic target in NSCLC [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B06.
Abstract LB-B07: Photoreceptor phosphodiesterase 6H expression regulates HCT116 proliferation and cell cycle progression
Authors: Yalaz Ceren , Haider Syed , Chen Jianzhou , Prevo Remko , Woodcock Mick , Harris Adrian L. , Muschel Ruth J.
Published: August 2018

Phosphodiesterase 6 (PDE6) protein complex, which is responsible for the turnover of the second messenger cyclic guanosine monophosphate (cGMP) and is involved in visual signal transduction of light stimuli, is highly expressed in certain cancer cell lines. Despite having been discovered in the outer segment of rods and cones of the retina, PDE6 protein is also detected in brain, adrenal glands, lungs, and gonads. Genes encoding the catalytic subunits of the enzyme are known to be upregulated in certain breast, non-small cell lung cancer and medulloblastoma cells. PDE6H, which encodes the cone-specific inhibitory gamma subunit of PDE6 protein, is part of the gene signature of achromatopsia and congenital stationary night blindness while its relation to cancer remains unclear. We recently identified PDE6H as a cell cycle controller in HCT116 colorectal cancer cell line, as part of an siRNA screen in which DNA content measurement via flow cytometry was undertaken to monitor cell cycle distribution. The screen was conducted with a custom library of metabolic genes commonly amplified in primary tumours, as well as those known to be involved in the key regulatory points of metabolic pathways.

We have shown that PDE6H knockdown and pharmacological inhibition of PDE6 protein hindered the activity of the protein complex, causing an accumulation of cGMP in various colorectal and breast cancer cell lines. This inhibition resulted in a G1 cell cycle arrest and disruption of G1/S checkpoint mechanism. PDE6H knockdown inhibited cell proliferation and cellular oxidative phosphorylation. It also altered mitochondrial morphology and induced apoptosis. Our results suggest that the gamma subunit is important for the function of photoreceptor phosphodiesterase PDE6 complex and PDE6H inhibition can be explored as an anti-proliferative strategy.

Citation Format: Ceren Yalaz, Syed Haider, Jianzhou Chen, Remko Prevo, Mick Woodcock, Adrian L. Harris, Ruth J. Muschel. Photoreceptor phosphodiesterase 6H expression regulates HCT116 proliferation and cell cycle progression [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B07.
Abstract LB-B09: Application of Fluorescence Correlation Spectroscopy as a novel tool to quantify target occupancy in cells and tumor tissue
Authors: Becker Frank , Hansen Kerrin , Crochiere Marsha , Landesmann Yosef , Hannus Stefan
Published: August 2018

The quantification of Target Occupancy (TO) in cells after application of an inhibitor is an important parameter to correlate the biochemical features of a drug with its in vivo efficacy. Experimental evidence of TO is a robust support of the target hypothesis and helps to identify optimal dosing of the drug. The sensitive correlation spectroscopy methods (FCS and FCCS) have been used successfully in the past to analyze drug- target interactions. The technology is fast, specific and allows detection of drug target interactions in smallest sample volumes. Here we adopted the FCCS method to quantify the occupancy of the XPO1 target in cells and tumors treated with the Selective Inhibitor of Nuclear Export (SINE) compound Selinexor (KPT-330), currently in advanced clinical trials in patients with hematological and solid cancers. We used a fluorescently labeled tracer (Leptomycin B -Cy5; LMB-Cy5) to quantify the loading of XPO1 with Selinexor and confirmed target specificity of the signal using a fluorescent labeled antibody detecting native XPO1 protein. The amount of Antibody-XPO1-tracer complex was determined by FCCS. Occupancy of XPO1 target with Selinexor decreases the amount of tracer-target complexes in a dose and time dependent manner. Target occupancy studies of XPO1 were carried out in different cancer relevant cell lines and xenograft tumors retrieved from treated animals. As such the approach can be applied to study target occupancy in vitro and in vivo.

Citation Format: Frank Becker, Kerrin Hansen, Marsha Crochiere, Yosef Landesmann, Stefan Hannus. Application of Fluorescence Correlation Spectroscopy as a novel tool to quantify target occupancy in cells and tumor tissue [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B09.
Abstract LB-B26: Novel eIF4A inhibitors target both cellular and microenvironment components in pancreatic ductal adenocarcinoma
Authors: Safari Maryam , Kim Yu Ri , Deng Changchun , Bates Susan E.
Published: August 2018

We are investigating the potential utility of targeting protein translation via inhibiting the translation initiation factor 4A (eIF4A) with a new class of Pateamine A derivatives to treat pancreatic ductal adenocarcinoma (PDAC). PDAC is one of the most refractory human malignancies with poor response to therapy and low survival rates. PDAC resistance to therapy is thought to comprise both cellular and microenvironment components. A key cellular aberration is thought to be the activating KRAS mutations found in over 90% of pancreatic cancers, signaling proliferation and survival. One postulated effect of KRAS mutation is to activate C-Myc, a central oncogene in PDAC, with elevated expression and high protein levels in over 40% of cases. A key microenvironment component is elevated hyaluronic acid content in the stroma of pancreatic cancers (4 to 12 times that of normal pancreatic tissues), which increases interstitial fluid pressure and is thought to impair drug perfusion efficiency, as well as the infiltration of immune cells. Here we report a promising therapeutic intervention for PDAC via targeting the eIF4A component of the translation initiation complex using a new class of Pateamine A derivatives, DMDA PatA and MZ735. Using cytotoxicity assays, immunoblotting, immunocytochemistry and confocal imaging we show activity of the Pateamine A derivatives on PDAC cell lines. Our primary data shows treatment with nanomolar concentrations of these compounds inhibits cell proliferation with IC50s ranging from 20 to 40nM for DMDAPatA and 5 to 20 nM for MZ735 in various PDAC cells. Immunoblotting shows a reduction in c-MYC, hyaluronic acid synthase HAS3, and hyaluronic acid receptor CD44 in PDAC cell lines following treatment with eIF4A targeting compounds. Finally, confocal imaging confirmed the reduction of hyaluronic acid and CD44 in the eIF4A inhibitor-treated PDAC cells. We believe Pateamine A derivatives represent a new class of anticancer agents that have the potential to address both altered cellular and microenvironment components in PDAC.

Citation Format: Maryam Safari, Yu Ri Kim, Changchun Deng, Susan E. Bates. Novel eIF4A inhibitors target both cellular and microenvironment components in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B26.
Abstract LB-B28: Abietane natural products as novel therapeutics against solid tumors
Authors: Lang Walter , Ling Taotao , Lee Jangsoon , Ueno Naoto T. , Rivas Fatima
Published: August 2018

Dysregulation of protein synthesis can lead to aberrant cellular proliferation and survival, a hallmark of cancer. Protein synthesis is a multistep process tightly controlled by specific translation factors. In eukaryotes, at the initiation point, the 43S preinitiation complex is formed by the association of several factors [eukaryotic initiation factor 2 (eIF2), GTP and methionyl initiator tRNA] with the 40S ribosomal subunit and the eukaryotic initiation factor 3 (eIF3). Over expression of eIFs has been reported in breast cancer, particularly in the most aggressive subtype, triple negative breast cancer (TNBC). More than one million global cases of breast cancer are diagnosed yearly and approximately 15% are characterized as triple negative breast cancer (TNBC), a subtype that lacks effective targeted therapy modalities. Recently, we generated a library of cell permeable abietane natural product derivatives (i.e. SJ6038) and showed that these compounds are more potent against TNBC over the estrogen receptor positive cellular models. These compounds were tagged with biotin for affinity enrichment and with a fluorescent chromophore to facilitate protein target(s) identification. A SILAC proteomics approach was used to distinguish specific binding targets from the non-specific ones. Gene Ontology analysis showed the targets identified were affiliated with regulation of protein synthesis. Ingenuity Pathway Analysis suggested that abietanes may exert their anticancer effects on critical biological pathways such as the eIF2, eIF4/p70S6K, and mTOR signaling pathways. Live cell experiments utilizing fluorescent abietane derivatives demonstrate that these probes localize to subsections of the ER, supporting the SILAC findings. Time of Exposure and Dose Response experiments further demonstrate that Abietane derivatives affect the eIF2 signaling axis. Using chemical biology and medicinal chemistry approaches, we show that abietane natural products preferentially promote cell death in TNBC cellular models by targeting the eIF2 signaling axis. These compounds will serve as unique chemical tools targeting the eIF2/eIF4 signaling axis and advance from hit- to lead for further preclinical development as a targeted therapy against aggressive solid tumors.

Citation Format: Walter Lang, Taotao Ling, Jangsoon Lee, Naoto T. Ueno, Fatima Rivas. Abietane natural products as novel therapeutics against solid tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B28.
Abstract LB-B34: LIF as a novel cancer immunotherapy target: modulating the tumor microenvironment with MSC-1, a humanized anti-LIF monoclonal antibody
Authors: Magram Jeanne , Pandya Naimish , Hoffman Kimberly , Hallett Robin , Giblin Patricia , Sinclair Angus , Anido Judit , Ruano Isabel Huber , Sala Ada , Pascual Monica , Chiganças Vanessa , Wasserman Robert , Seoane Joan
Published: August 2018

Leukemia Inhibitory Factor (LIF) is a pleiotropic cytokine involved in many physiological and pathological processes. LIF is highly expressed in a subset of tumors across multiple tumor types and has been shown to correlate with poor prognosis. LIF is hypothesized to contribute to tumor growth and progression by acting on multiple aspects of cancer biology, including immunosuppression of the tumor microenvironment and is a key regulator of cancer initiating cells (CICs). MSC-1, a first-in-class, humanized monoclonal antibody (IgG1), is a potent and selective inhibitor of LIF. MSC-1 leads to STAT3 inhibition by disrupting LIF signaling through the LIF receptor (LIFR). Blocking LIF with MSC-1 decreased tumor growth in multiple mouse tumor models and drove reprogramming of the tumor microenvironment through modulation of immunosuppressive macrophages and of several immune cell types. These findings form the basis of a robust therapeutic hypothesis, whereby MSC-1 treatment may lead to clinical activity in multiple cancer indications. Clinical testing is planned to initiate in the end of 2017 and trials will incorporate target engagement and PD biomarkers as well as efficacy endpoints.

Citation Format: Jeanne Magram, Naimish Pandya, Kimberly Hoffman, Robin Hallett, Patricia Giblin, Angus Sinclair, Judit Anido, Isabel Huber Ruano, Ada Sala, Monica Pascual, Vanessa Chiganças, Robert Wasserman, Joan Seoane. LIF as a novel cancer immunotherapy target: modulating the tumor microenvironment with MSC-1, a humanized anti-LIF monoclonal antibody [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B34.
Abstract 1001: Gut microbiota modulate T cell trafficking into human colorectal cancer
Authors: Cremonesi Eleonora , Governa Valeria , Garzon Jesus , Mele Valentina , Amicarella Francesca , Muraro Manuele G. , Droeser Raoul , Bolli Martin , Slotta-Huspenina Julia , Terracciano Luigi , Zajac Paul , Spagnoli Giulio C. , Eppenberger-Castori Serenella , Janssen Klaus-Peter , Borsig Lubor , Iezzi Giandomenica
Published: August 2018

Introduction: Colorectal cancer (CRC) is a leading cause of cancer-related death. CRC infiltration by immune cells, including cytotoxic CD8+ T cells (CTLs), IFN-gamma-producing T-helper 1 cells (Th1), Foxp3+ regulatory T cells (Tregs) and CD16+ MPO+ neutrophils, is associated with favorable prognosis. However, chemokines driving these cell populations into the tumor site, their cellular sources and their microenvironmental triggers remain to be elucidated. Aim: We investigated the chemokine/chemokine receptor network promoting CRC infiltration by immune cells associated to favorable prognosis. Results: CRC infiltration by immune cells was associated with defined chemokine gene signatures, including CCL5, CXCL9 and CXCL10 for cytotoxic T lymphocytes and T-helper 1 cells, and CCL17, CCL22 and CXCL12 for T-helper 1 and regulatory T cells. Most of these chemokine genes were expressed by tumor cells upon exposure to gut bacteria in vitro and in vivo. Indeed, chemokine expression levels were significantly higher in orthotopic xenografts than in intraperitoneal tumors, and were drastically reduced by antibiotic treatment of tumor-bearing mice. Importantly, in human CRC samples, extents of chemokine production and immune cell infiltration was significantly associated with bacterial loads. Conclusion: Gut microbiota stimulates chemokine production by CRC cells, thus favoring T cell recruitment into tumor tissues.

Citation Format: Eleonora Cremonesi, Valeria Governa, Jesus Garzon, Valentina Mele, Francesca Amicarella, Manuele G. Muraro, Raoul Droeser, Martin Bolli, Julia Slotta-Huspenina, Luigi Terracciano, Paul Zajac, Giulio C. Spagnoli, Serenella Eppenberger-Castori, Klaus-Peter Janssen, Lubor Borsig, Giandomenica Iezzi. Gut microbiota modulate T cell trafficking into human 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 1001.
Abstract 1002: Dysfunction of HPV16-specific CD8+ T cells derived from oropharyngeal tumors is related to the expression of Tim-3 but not PD-1
Authors: Partlova Simona , Hladikova Kamila , Koucky Vladimir , Fialova Anna , Spisek Radek , Boucek Jan , Zabrodsky Michal , Halaska Michael , Tachezy Ruth , Grega Marek , Fonteneau Jean-François
Published: August 2018

Human papillomavirus (HPV) type 16 infection is one of the most important etiological agents of oropharyngeal squamous cell carcinoma. Patients with HPV-positive squamous cell carcinomas of the head and neck were reported to have a better clinical outcome than patients with HPV-negative tumors. Because HPV16 E6 and E7 oncoproteins are highly immunogenic and constitutively expressed within the tumor cells, HPV-specific T cell immunity may contribute to the better prognosis of HPV-positive tumors. We analyzed the frequency, phenotype and function of HPV16 E6/E7-specific tumor-infiltrating T cells (TILs) in oropharyngeal tumors and tested the effect of anti-PD1 mAb (nivolumab), soluble Tim-3 (sTim-3) and homeostatic in vitro expansion on these characteristics. We show that 73.1% of HPV-associated oropharyngeal tumors are positive for HPV16-specific T cells capable of producing IFNγ upon stimulation with E6/E7 peptide pools. These IFNγ-producing HPV-specific TILs were mainly PD-1+Tim-3-CD8+ T cells, identifying Tim-3 rather than PD-1 as a marker of T cell dysfunction. Consequently, specific IFNγ production was further enhanced by combined nivolumab plus sTim-3 treatment, but not with nivolumab alone. Our data provide the rationale for exploring additional setups of anti-PD-1 mAb treatment, including combinations with other check-point blockers, such as Tim-3 and/or with HPV16-directed therapeutic vaccines.

Citation Format: Simona Partlova, Kamila Hladikova, Vladimir Koucky, Anna Fialova, Radek Spisek, Jan Boucek, Michal Zabrodsky, Michael Halaska, Ruth Tachezy, Marek Grega, Jean-François Fonteneau. Dysfunction of HPV16-specific CD8+ T cells derived from oropharyngeal tumors is related to the expression of Tim-3 but not PD-1 [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 1002.
Abstract 1020: Characterization of the immune landscape and analysis of tumor response after anti-PD-1 blockade in a 3D ex vivo system of non-small lung cancer
Authors: Mediavilla-Varela Melanie , Page Melba Marie , Kreahling Jenny , Altiok Soner
Published: August 2018

Background: Immuno-oncology drugs face the challenge that cell lines and animal models cannot recapitulate the full intimacies of the tumor microenvironment. Although these models have aided in the discovery of new medical modalities, only 8% of these models have had a successful translation. Furthermore, the phenotype of the microenvironment of various tumor types has not been assessed in depth. Nilogen Oncosystems' proprietary 3D-EX vivo drug screening platform analyzes fresh patient tumor tissue that remains embedded in its natural environment. With our model, the characteristics of the tumor microenvironment can be accurately revealed in response to checkpoint inhibitors in non-small cell lung cancer (NSCLC) and correlates can be found among different platforms.

Methods: For the 3D-EX vivo platform, microspheres were produced from procured fresh tumor tissue from NSCLC cancer patients. They were then treated with Keytruda ex vivo and treatment-mediated changes in TIL subpopulations were analyzed using flow cytometric analysis, cytokine release by Bio-Rad's 17-plex cytokine assay as well as gene expression by NanoString's 770 gene Immune Panel.

Results: Ex vivo treatment of the 3D microspheres with Keytruda, showed significant changes in T-cell activation and immune cell populations in 26% of NSCLC tumors. This was observed by the simultaneous increase in IFN-γ and TNF-α upon cytokine analysis. Furthermore, we found a differential expression of signature genes such as CD8, CXCL10, CXCL9, EOMES, Granzyme A/B, IFN-γ, related to T-cell subpopulations via Nanostring analysis, which was accompanied by an increase in Granzyme B via flow cytometry.

Conclusion: The positive and negative associations between expression of immune function genes, TIL activation, and cytokine production by ex vivo treatment shows that Nilogen Oncosystems' 3D-Ex vivo platform can accurately recapitulate the tumor microenvironment and exhibits the importance of a comprehensive analysis of the tumor immune microenvironment for a better understanding of the mechanism of action of immuno-oncology drugs that may aid in developing biomarkers that can be used for patient selection.

Citation Format: Melanie Mediavilla-Varela, Melba Marie Page, Jenny Kreahling, Soner Altiok. Characterization of the immune landscape and analysis of tumor response after anti-PD-1 blockade in a 3D ex vivo system of non-small lung 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 1020.
Abstract 106: Shock and ROR! Targeting ROR1 and ROR2 in a preclinical patient-derived model of ovarian cancer
Authors: Henry Claire E. , Hacker Neville F. , Ford Caroline E.
Published: August 2018

BACKGROUND: New targets for ovarian cancer treatment are critically needed. The Wnt receptors ROR1 and ROR2 are overexpressed in all histotypes of ovarian cancer and appear to play a role in both the tumour and the surrounding microenvironment. We have previously conducted an extensive suite of in vitro experiments, exploring the functional role of ROR1 and ROR2 in ovarian cancer. Silencing either receptor using siRNA inhibited ovarian cancer cell migration and invasion, and silencing both receptors had an even stronger inhibitory effect on the metastatic potential of ovarian cancer cellS. We have also shown that silencing ROR1 and ROR2 sensitises chemoresistant ovarian cancer cells to chemotherapy. Combined, these studies have confirmed the importance of RORs in ovarian cancer, and provided a strong argument for these receptors potential as clinical targets.

However, these previous studies have utilised simple 2D in vitro models to investigate cancer cell growth and migration, which does not allow investigation of stromal involvement in ROR driven metastasis.

AIM: To investigate targeting ROR1 and ROR2 in a 3D primary co-culture model of epithelial ovarian cancer dissemination to the omentum.

METHODS: Primary fibroblasts (NOF) and mesothelial (HPMC) cells were isolated from fresh samples of omentum collected from women with benign or non-metastatic conditions and cultured with collagen to produce a organotypic 3D model. Stable shRNA knockdown of ROR1, ROR2 and double ROR1/ROR2 in OVCAR4 ovarian cancer cells were incorporated into the 3D model to measure adhesion, or using a transwell to measure invasion. Gene expression changes in primary cells upon OVCAR4 interaction was evaluated using indirect transwell co-culture.

RESULTS: Double knockdown of ROR1 and ROR2 strongly inhibited cell adhesion (p<0.05) and invasion (P<0.05) to the omentum model. ROR2, but not ROR1, was up- regulated in primary fibroblasts when cultured with OVCAR4 (P=0.05) and ectopic overexpression of ROR2 in NOFs increased cell migration.

CONCLUSION: The combination of ROR1 and ROR2 signalling influences ovarian cancer dissemination to the omentum, however ROR2 may also play a specific role in stromal activation during metastasis. Therefore, targeting both ROR1 and ROR2 may be a powerful approach to treating ovarian cancer. The development of a number of monoclonal antibodies targeting ROR1 currently in phase 1 trials for other tumour types makes this clinically feasible in the near future.

Citation Format: Claire E. Henry, Neville F. Hacker, Caroline E. Ford. Shock and ROR! Targeting ROR1 and ROR2 in a preclinical patient-derived model of ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 106.
Abstract 1081: TCF12 regulates metastasis-promoting genes in pancreatic cancer
Authors: Maurin Michelle M. , Newberg Justin Y. , Black Mik A. , Jones Devin J. , Jenkins Nancy A. , Copeland Neal G. , Mann Karen M.
Published: August 2018

Pancreatic ductal adenocarcinoma (PDAC) is a highly metastatic disease with poor patient outcome. While acquired oncogenic activation of KRAS is an initiating event, disease progression requires functional perturbation of additional genes and cellular pathways. We have taken a comparative genomics approach to identify progression drivers using a mouse model of metastatic PDAC driven by Sleeping Beauty (SB) insertional mutagenesis in the context of oncogenic Kras (SB PDAC). Statistical enrichment analysis of high read depth insertions from SB PDAC tumors defined 48 drivers predicted to contribute to disease progression. Among these drivers, Tcf12 was associated with increased metastatic incidence in the SB PDAC mouse model. We demonstrated significant increases in primary tumor burden and metastatic spread by single gene knockdown of TCF12 in an orthotopic mouse model of pancreatic cancer. TCF12 encodes a basic helix-loop-helix (bHLH) transcription factor that partners with other bHLH transcription factors in different cellular contexts, including TWIST, a transcription factor involved in epithelial-to-mesenchymal transition (EMT), PTF1A, a pancreas-specific transcription factor with essential roles in pancreas development and cancer, and SMAD3, a TGFß-related transcription factor involved in cellular migration. TCF12 resides in a chromosomal region (15q21) frequently lost in pancreatic cancer, and decreased TCF12 expression is associated with poor patient prognosis. RNASeq analysis identified genes with TCF12 consensus binding sites within 10 kb of their promoter regions that are differentially expressed upon TCF12 knockdown. A subset of these genes function in Wnt or TGF-beta signaling pathways and show consistent decreases in expression upon TCF12 knockdown in orthotopic tumors. ChipSeq analysis and functional validation studies are ongoing to confirm these genes are direct TCF12 transcriptional targets involved in pancreatic tumor metastasis.

Citation Format: Michelle M. Maurin, Justin Y. Newberg, Mik A. Black, Devin J. Jones, Nancy A. Jenkins, Neal G. Copeland, Karen M. Mann. TCF12 regulates metastasis-promoting genes in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1081.