bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3490: Mechanism of action for antioxidant Twendee X on hepatic cancer cell line HepG2
Authors: Inufusa Haruhiko
Published: August 2018

[Purposes of the study] Reactive oxygen species (ROS) are associated with clinical condition of cancer patient, and closely involved in cancer growth and metastasis. Twendee X (TWX) is a composition consisting of Vitamin C, Glutamine, Cystine, Riboflavin, Succinic acid, Fumaric acid, Coenzyme Q10, and Niacin. TWX strongly reduces oxygen radicals or ROS in vitro and in vivo (World Intellectual Property Organization WO2013/072441 A1 COMPOSITION FOR PROTECTION AGAINST CELL-DAMAGING EFFECTS). TWX has significant effects of reducing tumor growth and inhibition metastasis. Hydrogen peroxide level (d-ROMs test) of tumor bearing nude mouse in serum was significantly reduced and increased NK-activity (2016 AACR meeting). Mechanism of action was examined for TWX using hepatic cancer cell line HepG2.

[Experimental procedures] Human hepatic cancer cell line HepG2 was used in this study. Cell proliferation experiments of antioxidant by ICDD (France). Metabolome analysis was by Human Metabolome Technologies (Tokyo). In both experiments, HepG2 cell line treated one hour with TWX (60ug/ml). [Data] TWX inhibits the RAF-MEK-ERK pathway starting from ERK. Metabolome analysis shows total shift of metabolism, and especially increase mitochondrial metabolism and ATP production. [Conclusion] Effects of TWX on cancer cell line are not only increase immune activity, but also inhibit proliferation pathway, and mitochondrial metabolism change. Several publications using Vitamin-E or Cysteine as anti-oxidant showed cancer progression or increase metastasis. However, Vitamin-E or Cysteine has limited anti-oxidant effects and hydrogen peroxide level (d-ROMS test) dose not change. On the other hand, TWX has significant effects reduce hydrogen peroxide level, and Japanese Association for Dementia Prevention already started clinical trial for MCI and after stroke patients. For the cancer patients, clinical trial using TWX combination with standard therapy will be start soon.

Citation Format: Haruhiko Inufusa. Mechanism of action for antioxidant Twendee X on hepatic cancer cell line HepG2 [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 3490.
Abstract 3514: Differential metabolic targeting of PDAC cells with Zaprinast
Authors: Rao Yi , Gammon Seth , Piwnica-Worms David
Published: August 2018

Many pancreatic cancers consume excessive glutamine as a preferable source for anaplerosis, maintenance of proper redox state, and synthesis of intermediates for cancer cell growth and proliferation. Abrogating glutamine metabolism is under investigation to inhibit the proliferation/ growth of glutamine-addicted cancers. In addition, inhibition of the mitochondrial pyruvate carrier (MPC1) can potentially induce cancer cells to become more dependent on glutaminolysis. In this regard, Zaprinast, originally developed as a phosphodiesterase 5 (PDE5) inhibitor, has recently been identified as an inhibitor of both glutaminase as well as MPC1. We compared the growth inhibition effect of Zaprinast against BPTES, a targeted glutaminase inhibitor, and UK5099, a targeted MPC1 inhibitor, as well as the combination of BPTES and UK5099. Using pancreatic ductal adenocarcinoma (PDAC) cells, KRAS mutant (Mia Paca2) and KRAS wildtype (Panc03.27), KRAS mutant neuroendocrine (QGP1), and normal pancreatic epithelial cells (HPNE), Zaprinast alone showed more potent growth inhibition in all tested cells lines than single agent treatment with BPTES or UK5099. Further, Zaprinast effectively depleted mitochondrial reserve respiratory capacity similar to UK5099, rendering mitochondria more vulnerable to oxidative stress or pharmacological inhibitors targeting the electron transport chain. In addition, Zaprinast treatment of cancer cells differentially elevated ROS levels greater than 10-fold compared to normal HPNE cells (ROS levels: Mia Paca2 ~ Panc03.27 > QGP1 > HPNE). However, the increase in ROS levels did not correlate with Zaprinast-induced growth inhibition (IC50: Mia Paca2=300 µM, Panc03.27=110 µM, HPNE=650 µM, QGP1 >1 mM). Although Zaprinast-induced growth inhibition was selective toward adenocarcinoma cells but not the neuroendocrine subtype, the selective Zaprinast-induced elevation of ROS levels toward all cancer subtypes renders possibilities for potential synthetic lethality when combined with other metabolic inhibitors.

Citation Format: Yi Rao, Seth Gammon, David Piwnica-Worms. Differential metabolic targeting of PDAC cells with Zaprinast [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 3514.
Abstract 382: Inhibition of mitochondrial spare respiratory capacity by new securinine derivatives as a novel differentiation therapy for acute myeloid leukemia
Authors: Karunanithi Sheela , Liu Ruifu , Roe Anne , Moreton Stephen , Hou Yongchun , Oldford Natasha , Goyco Juan Valentin , Wald David
Published: August 2018

Acute myeloid leukemia (AML) is characterized by a differentiation block in myeloid progenitors resulting in aberrant proliferation of immature myeloid cells. Therapeutics that induce differentiation of leukemic cells would overcome the high toxicity and low potency of conventional chemotherapy. Securinine, a plant-derived alkaloid from Securinega suffruticosa, has been shown to be a promising AML differentiation agent through an unclear mechanism, but is limited by its low potency (~20μM) and toxicity. We performed lead optimization chemistry on securinine that led to derivatives 250 and 317, with nanomolar efficacy and reduced toxicity. These derivatives effectively inhibit growth of AML cell lines and patient samples. Treatment of AML cell lines with compounds 250 and 317 induces the expression of differentiation markers CD11b and CD14c. Compounds 250 and 317 significantly reduced leukemic burden in a circulating model of AML in NSG mice. Additionally, these compounds were effective in inhibiting cell growth of HL60 cells that are resistant to a conventional chemotherapeutic, doxorubicin. By using the drug-affinity responsive target sensitivity (DARTS) assay, we identified that the redox mediator thioredoxin reductase (TrxR) is the primary target of these derivatives. 250 and 317 inhibit TrxR activity in vitro and in cells, and the resultant alterations in redox status are overcome by addition of a scavenger of reactive oxygen species (ROS), N-acetylcysteine. The ensuing oxidative metabolic stress rapidly alters the metabolite profile in AML cells including depletion of malate. Securinine derivatives decreased basal oxygen consumption rate and spare respiratory capacity, resulting in differentiation and cell death. Downregulating TrxR1 expression increased the sensitivity of OCI-AML3 cells to compounds 250 and 317. Together, we show here that these new derivatives of securinine exhibit preclinical efficacy in AML by decreasing mitochondrial spare respiratory capacity through the inhibition of TrxR.

Citation Format: Sheela Karunanithi, Ruifu Liu, Anne Roe, Stephen Moreton, Yongchun Hou, Natasha Oldford, Juan Valentin Goyco, David Wald. Inhibition of mitochondrial spare respiratory capacity by new securinine derivatives as a novel differentiation therapy for acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 382.
Abstract 3833: SEA-BCMA: A highly active enhanced antibody for multiple myeloma
Authors: Epps Heather Van , Anderson Martha , Yu Changpu , Klussman Kerry , Westendorf Lori , Carosino Chris , Manlove Luke , Cochran Julia , Neale Jason , Benjamin Dennis , Ryan Maureen
Published: August 2018

Despite a number of new therapies for multiple myeloma (MM) most patients relapse, require multiple lines of therapy, and ultimately succumb to disease. Therapies that are well tolerated and active remain an unmet need. Currently, successful MM therapies combine agents with different mechanisms of action and safety profiles. Therapeutic antibodies have recently altered the MM treatment paradigm, allowing patients to achieve deeper responses with minimal added toxicity. SEA-BCMA is a humanized afucosylated IgG1 antibody targeting BCMA, which shows preclinical evidence of encouraging activity and tolerability. BCMA is expressed at the surface of plasma cells, and induces proliferative signals through the binding of its ligands APRIL and BAFF. SEA-BCMA acts through three mechanisms of action. One, it engages in increased binding to FcγRIII through SEA technology leading to enhanced antibody dependent cellular cytotoxicity. Two, it mediates antibody dependent cellular phagocytosis. Three, it blocks the proliferative signals from BCMA ligand binding. This antibody shows activity in all seven tumor xenograft models tested, inducing tumor delays at doses as low as 0.1mg/kg, and generating prolonged survival and durable regressions with repeat dosing. SEA-BCMA is active on tumor xenografts expressing as few as 2000 surface copies of BCMA antigen. Both effector function and ligand blocking contribute to overall in vivo activity. In the absence of effector cell recruitment, ligand blocking alone can induce prolonged durable regressions. In addition, SEA-BCMA can target MM cells in the presence of soluble BCMA. SEA-BCMA is tolerated up to 100mg/kg in cynomolgus monkey, reflecting lack of toxicity from cellular interactions with the Fc portion of the Ab, since SEA-BCMA does not bind to the cynomolgus monkey antigen. SEA-BCMA displays a 12-day half-life in these animals. In vitro testing with human PBMCs or bone marrow mononuclear cells induced minimal cytokine production in the presence of BCMA target. In summary, SEA-BCMA is highly active and well tolerated in preclinical models and is a strong candidate for treatment of MM patients.

Citation Format: Heather Van Epps, Martha Anderson, Changpu Yu, Kerry Klussman, Lori Westendorf, Chris Carosino, Luke Manlove, Julia Cochran, Jason Neale, Dennis Benjamin, Maureen Ryan. SEA-BCMA: A highly active enhanced antibody for multiple myeloma [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 3833.
Abstract 3946: Androgen receptor drives differential gene expression in KRAS-mediated non-small cell lung cancer
Authors: Wang Albert Roy , Beyer Hope , Brennan Sean , Stiles Shannon , Wiese Dylan , Buehler Darya , Saeed Anwaar , Baschnagel Andrew M. , Iyer Gopal
Published: August 2018

The Cancer Genome Atlas (TCGA) has identified alterations such as amplification, deletion, or mutation of androgen receptor (AR) in about 5% of human lung squamous cell carcinoma and adenocarcinoma. The expression of AR in human lung appears to play a crucial role in lung development and type II pneumocytes (PTII) maturation, but unlike in prostate cancer, the role of AR is not well known in non-small cell lung cancer (NSCLC). In our study, we demonstrated AR in NSCLC cell lines translocated into the nucleus over time when stimulated with synthetic androgen R1881, while addition of enzalutamide (MDV3100) or AR siRNA reduced AR nuclear localization. Using an NSCLC human tissue microarray also revealed that 10 out of 88 patients (11%) have positive AR immunohistochemical staining. Preliminary data from clonogenic assay indicated that enzalutamide might have radiosensitizing effect on certain NSCLC cells at 2 Gy, further suggesting the involvement of AR in lung cancer and its potential therapeutic value as a target. With the induction of R1881 for 30 minutes to 24 hours, certain NSCLC cells display decreased AR mRNA (0.5-0.8 fold), while others show modest increase (up to 1.5 fold), suggesting that AR regulation in these cells might be different due to their mutational landscapes. We then used predesigned 384 well panels from Bio-Rad to survey the effects of R1881, enzalutamide, and AR siRNA on mRNA expression levels of selected NSCLC cells. Distinct expression profiles were observed between cells that have wild-type and mutated KRAS. Taken together, these findings suggest that the AR signaling could be different based on KRAS mutational profiles of NSCLCs, and further work is required to reveal the underlying mechanism. Chromatin immunoprecipitation (ChIP) assay and RNA sequencing will be utilized to investigate AR interactions with androgen response elements and differential expressions in KRAS-driven NSCLC cells, respectively.

Citation Format: Albert Roy Wang, Hope Beyer, Sean Brennan, Shannon Stiles, Dylan Wiese, Darya Buehler, Anwaar Saeed, Andrew M. Baschnagel, Gopal Iyer. Androgen receptor drives differential gene expression in KRAS-mediated non-small cell 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 3946.
Abstract 3967: Potent treatment combinations with CIB1 depletion in triple-negative breast cancer
Authors: Chung Alexander H. , Leisner Tina M. , Dardis Gabrielle , Bivins Marissa , Parise Leslie V.
Published: August 2018

The current standard of care for triple-negative breast cancer (TNBC) patients is limited to chemotherapy, radiation, and/or surgery. Therefore, there is a pressing need for new targeted therapies to improve clinical efficacy, reduce toxicity, and overcome resistance. We previously found that CIB1 depletion induces cell death selectively in TNBC cells, while sparing normal cells. The purpose of this study was to ask whether depleting CIB1 further enhances tumor-specific killing when combined with either the commonly used chemotherapeutic, docetaxel, or the cell death-inducing ligand, TRAIL. To address this, we targeted CIB1 by RNA interference in MDA-MB-436, MDA-MB-231, MDA-MB-468 TNBC cells and ME16C normal breast epithelial cells alone or combination with docetaxel or TRAIL. Cell death was quantified via trypan blue exclusion using FACS and cell death mechanisms were analyzed by Western blotting. We found that the combination of CIB1 depletion with docetaxel significantly enhanced tumor-specific cell death relative to each treatment alone. The enhanced cell death strongly correlated with caspase-8 activation, a hallmark of death receptor-mediated apoptosis. Because CIB1 depletion also upregulated the death receptor, TRAIL-R2, we tested for sensitization to TRAIL. While TRAIL alone had no effect on cell viability, CIB1 depletion profoundly sensitized tumor cells to TRAIL, resulting in a synergistic increase in cell death. In addition to death receptor-mediated apoptosis, both combination treatments activated a non-apoptotic mechanism, called paraptosis. Interestingly, these combination treatments also induced nearly complete death of docetaxel-resistant MDA-MB-436 cells, again via apoptosis and paraptosis. In contrast, neither combination treatment induced cell death in normal ME16C cells. In conclusion, our results demonstrate that the novel combinations of CIB1 depletion with docetaxel or TRAIL significantly enhance naive and docetaxel-resistant TNBC cell death via increased death receptor-mediated apoptosis and paraptosis, suggesting that targeting CIB1 may also overcome resistance to chemotherapeutics. Taken together, combination therapies that target CIB1 could prove to be a safe and durable strategy for treatment of TNBC and potentially other cancers.

Citation Format: Alexander H. Chung, Tina M. Leisner, Gabrielle Dardis, Marissa Bivins, Leslie V. Parise. Potent treatment combinations with CIB1 depletion in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3967.
Abstract 4068: Galectin-1 intensifies immunosuppression in head and neck cancer by boosting myeloid-derived suppressive cell (MDSC) expansion
Authors: Nambiar Dhanya K. , Aguilera Todd A. , Bloomstein Joshua Daniel , Jiang Dadi , Cao Hongbin , Koong Albert , Le Quynh Thu
Published: August 2018

Head and neck squamous cell carcinoma (HNSCC), with more than 450,000 newly diagnosed cases annually, is the 6th most common cancer worldwide. Recent literature shows that tumor induced suppression of the host immune system is critical to HNSCC progression and metastases. Tumor secreted factors directly influence the expansion of myeloid-derived suppressor cells (MDSCs), which are considered important contributors for tumor to escape from immune surveillance. The presence of expanded MDSCs peripherally and within the tumor microenvironment is associated with metastasis and recurrence after definitive treatment in HNSCC patients. In this study, we evaluated at the role of tumor secreted Galectin-1 in recruitment of MDSCs in HNSCC. We utilized HPV-negative as well as HPV-positive HNSCC models to test the effect of Galectin-1 knockout (KO) in the tumor microenvironment. CRISPR/Cas9 knock out Galectin-1 in MOC2 and MEERL models resulted in a ~50% reduction in the tumor growth. We further analyzed the tumor microenvironment of these tumors for MDSC recruitment. Both MOC2 and MEERL tumors showed highly inflammatory phenotype, with significant presence of MDSCs (>70% of CD45+ cells were of myeloid origin). Knocking out Galectin-1 led to a significant decrease in CD11b+Gr1+ MDSCs in both MOC2 and MEERL tumors. We also observed high level of MDSCs in both the spleen and the peripheral blood of mice bearing Galectin-1 wildtype (WT) tumors in both models. In contrast, knocking out Galectin-1 from the tumor reduced systemic MDSCs levels. To investigate how Galectin-1 modulates MDSC levels, we compared the secreted cytokine profiles between MOC2 WT and Galectin-1 KO tumors. Galectin-1 null cells secreted significantly lower levels of monocyte and macrophage recruiting chemokines: CCL5, CXCL1, and G-CSF. Analysis of the gene expression signatures of MOC-2 WT and KO tumor cells revealed critical signaling pathways differences, including down-regulation of PI3K-AKT pathway in KO-tumors. In conclusion, the present study reveals that tumor derived Galectin-1 can stimulate the recruitment of MDSCs, thereby potentiating an inflammatory tumor microenvironment which supports HNSCC progression. (Acknowledgements: BMS for Anti-Gal1 Abs. Dr R. Uppaluri, for MOC-2 oral cancer cells, Dr. W. Spanos for MEERL cells)

Citation Format: Dhanya K. Nambiar, Todd A. Aguilera, Joshua Daniel Bloomstein, Dadi Jiang, Hongbin Cao, Albert Koong, Quynh Thu Le. Galectin-1 intensifies immunosuppression in head and neck cancer by boosting myeloid-derived suppressive cell (MDSC) expansion [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 4068.
Abstract 432: Establishment of reference samples for the detection of somatic variants in cancer
Authors: Fang Li Tai , Xiao Wenming ,   Somatic Mutation Working Group, SEQC2 Consortium
Published: August 2018

The FDA is well aware of the crucial role that next generation sequencing (NGS) plays in precision medicine, and this technology is already leveraged in regulatory decision-making. However, lack of well-characterized and community-validated reference samples and data benchmarks creates a potential challenge for development and review of NGS applications.

To address this challenge, 60+ institutions and 160+ scientists have come together to form a somatic mutation working group within the FDA-led Sequencing Quality Control Phase II (SEQC2) consortium. We present genomic samples with high-confidence germline and somatic mutation data in order to establish a community standard against which new tools and platforms can be evaluated and benchmarked. Our initial reference samples are a triple-negative breast cancer cell line: HCC1395, and its B lymphocyte derived normal cell line: HCC1395BL. To minimize bias, we sequenced these two cell lines at up to 10 sequencing centers using multiple sequencing platforms. In totality, we have performed whole genome sequencing of the two cell lines to coverage depths of 2400X on Illumina HiSeq, 450X on NovaSeq, 600X on 10X Genomics and 50X on PacBio. Additionally, we performed whole exome sequencing at 1200X on HiSeq and 100X on Ion Torrent. Lastly, we sequenced (RNAseq) >5000 single cells for cellular heterogeneity studies. We applied machine learning algorithms to define high confidence mutation calls with datasets from 8 NGS platforms combined with 9 bioinformatics analysis pipelines. Each mutation in the “gold standard” dataset was annotated with confidence scores and supporting evidence. To independently evaluate accuracy of our call set, we selected a subset of mutation calls for additional validation using orthogonal methods, including ddPCR.

To summarize, we present the first release of high-confidence germline variants and somatic mutations on a pair of well characterized cell lines. We plan to continuously advance accuracy and completeness of our reference sample data, in order to serve as a standard for the community for years to come.

Citation Format: Li Tai Fang, Wenming Xiao, Somatic Mutation Working Group, SEQC2 Consortium. Establishment of reference samples for the detection of somatic variants in 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 432.
Abstract 4344: Low mitochondrial copy number induces resistance to chemotherapy in esophageal cancer
Authors: Yamada Moyuru , Tanaka Koji , Masuike Yasunori , Makino Tomoki , takahashi Tsuyoshi , Kurokawa Yukinori , Yamasaki Makoto , Nakajima Kiyokazu , Mori Masaki , Doki Yuichiro , Odagiri Kabuki
Published: August 2018

Introduction: Esophageal squamous cell carcinoma (ESCC) is the one of the deadliest gastrointestinal cancers. The important problem in treating ESCC is resistance to chemotherapy. A few previous reports showed that mitochondrial DNA (mtDNA) copy number was decreased in esophageal squamous cell carcinoma (ESCC). However, it is unclear whether alteration of mtDNA copy number affects the effect of chemotherapy for ESCC. In the present study, we investigated the relationship between mtDNA copy number and the effect of chemotherapy in ESCC.

Methods: To analyze the mtDNA copy number of surgically resected primary tumors from ESCC patients, qPCR for Cytochrome Oxidase I was performed. Human ESCC cells (TE8 and TE11) with decreased mtDNA copy number were established by knockdown of mitochondrial transcription factor A (TFAM). Those cells were treated with 5FU, cisplatin and docetaxel. The response of chemotherapy was accessed by cell viability assay (WST assay) and apoptosis assay (flow cytometry). To investigate the mechanism of resistance of chemotherapy, expression of EMT-related genes such as CDH1, CDH2, vimentin and zeb1 was measured by qPCR. Wound healing assay and invasion assay were performed to access the cell migration and invasion.

Results: Patients were categorized into higher and lower subgroups according to the median value of the mtDNA copy number. Lower mtDNA copy number group was significantly correlated with tumor depth and pathologic response of chemotherapy. ESCC patients with lower mtDNA copy number had significantly poorer 5-year recurrence-free and overall survival than higher group. By shRNA knockdown of TFAM gene expression, mtDNA copy number decreased to 40% in TE8 and 60% in TE11 compared with non-target control cells. Viability of mtDNA-depleted TE8 and TE11 treated with chemotherapy was higher than that of control cells. Apoptosis assay showed the percentages of apoptosis induced by chemotherapy in mtDNA-depleted TE8 and TE11 were lower than those of control cells. Invasion assay and wound healing migration assay showed that mtDNA-depleted TE8 and TE11 were significantly more invaded and migrated than the control-sh cells.

Conclusions: These results suggest that low mitochondrial copy number induces resistance to chemotherapy in esophageal cancer.

Citation Format: Moyuru Yamada, Koji Tanaka, Yasunori Masuike, Tomoki Makino, Tsuyoshi takahashi, Yukinori Kurokawa, Makoto Yamasaki, Kiyokazu Nakajima, Masaki Mori, Yuichiro Doki, Kabuki Odagiri. Low mitochondrial copy number induces resistance to chemotherapy in esophageal 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 4344.
Abstract 4413: DNA methylation of miR-7 is a mechanism involved in platinum response through MAFG overexpression in cancer cells
Authors: Vera Olga , Jimenez Julia , Rodriguez-Antolin Carlos , Pernia Olga , Rodriguez Carmen , Soto Javier , Rosas Rocio , Esteban-Rodriguez Isabel , Perona Rosario , Castro Javier de , Ibáñez-de-Cáceres Inmaculada
Published: August 2018

One of the major limitations associated with platinum use is the resistance that almost invariably develops in different tumor types. In the current study, we sought to identify epigenetically regulated microRNAs as novel biomarkers of platinum resistance in lung and ovarian cancers, the ones with highest ratios of associated chemo-resistance. Experimental procedures: We combined transcriptomic data from microRNA and mRNA under the influence of an epigenetic reactivation treatment in a panel of four paired cisplatin -sensitive and -resistant cell lines, followed by real-time expression and epigenetic validations for accurate candidate selection in 19 human cancer cell lines. To identify specific candidate genes under miRNA regulation, we assembled “in silico” miRNAs and mRNAs sequences by using ten different algorithms followed by qRT-PCR validation. Functional assays of site-directed mutagenesis and luciferase activity, miRNAs precursor overexpression, silencing by antago-miR and cell viability were performed to confirm their specificity in gene regulation. Results were further explored in 187 primary samples obtained from ovarian tumors and controls. Results: We identified 4 candidates, miR-7, miR-132, miR-335 and miR-148a, which deregulation seems to be a common event in the development of resistance to cisplatin in both tumor types. miR-7 presented specific methylation in resistant cell lines, and was associated with poorer prognosis in ovarian cancer patients. Our experimental results strongly support the direct regulation of MAFG through miR-7 and their involvement in the development of CDDP resistance in human tumor cells. Conclusion: The basal methylation status of miR-7 before treatment may be a potential clinical epigenetic biomarker, predictor of the chemotherapy outcome to CDDP in ovarian cancer patients. To the best of our knowledge, this is the first report linking the regulation of MAFG by miRNA-7 and its role in chemotherapy response to CDDP. Furthermore, this data highlights the possible role of MAFG as a novel therapeutic target for platinum resistant tumors.

Citation Format: Olga Vera, Julia Jimenez, Carlos Rodriguez-Antolin, Olga Pernia, Carmen Rodriguez, Javier Soto, Rocio Rosas, Isabel Esteban-Rodriguez, Rosario Perona, Javier de Castro, Inmaculada Ibáñez-de-Cáceres. DNA methylation of miR-7 is a mechanism involved in platinum response through MAFG overexpression in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4413.
Abstract 4425: PARP1 inhibition impedes Warburg effect in breast cancer cells through modulation of the miR-98/STAT3 signaling axis
Authors: Shukla Nidhi
Published: August 2018

Cancer cells have increased metabolic autonomy and rewired cellular metabolism in order to meet the bioenergetics, biosynthetic, and redox demands during tumorigenesis. They tend to depend on glycolysis rather than oxidative metabolism to meet their energy demand. Therefore, elucidation of the key regulators of metabolism could potentially aid in devising selective and promising anti-cancer therapies. MicroRNAs (miRNAs) have recently attained prominence as key regulators of cancer cell metabolism. MiRNAs are known to regulate metabolism in cancer cells either through the direct regulation of key metabolic enzymes, tumor suppressors and oncogenes or the indirect modulation of master transcription factors and signalling pathways. Therefore dysregulation of cellular metabolism by miRNAs contributes significantly to cancer progression. Current therapeutic strategies include: (a) targeting metabolic pathways that are strictly required for cancer cells, or (b) impairing an oncogenic signalling axis involved in cancer metabolic rewiring. The poly (ADP-ribose) polymerase (PARP) inhibitors, used in the treatment of DNA repair deficient tumours are also known to facilitate oxidative metabolism. However, the effect of PARP inhibition on glycolytic metabolism is yet to be elucidated. In the present study we aimed to identify novel miRNAs involved in the regulation of glycolytic metabolism after PARP1 inhibition. MiRNA microarray analysis revealed upregulation of the tumor suppressor miR-98 in PARP1 inhibitor treated MDA-MB-231 breast cancer cells. Further mechanistic studies confirmed miR-98 regulation of STAT3 expression and downregulation of key glycolytic enzymes HK2 and PKM2 leading to decreased Warburg effect. MiR-98 overexpression or PARP1 inhibition in combination with miR-98 antagomir showed downregulation of STAT3, HK2 and PKM2. Glucose consumption and HK2 activity were decreased upon PARP1 inhibition associated miR-98 expression. Collectively, these results showed that miR-98 is a critical regulator of PARP1 inhibition associated reduction of Warburg effect in breast cancer cells.

Citation Format: Nidhi Shukla. PARP1 inhibition impedes Warburg effect in breast cancer cells through modulation of the miR-98/STAT3 signaling axis [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 4425.
Abstract 4548: Host immunogenetics and hyperprogression under PD1/PD-L1 checkpoint inhibitors
Authors: Refae Sadal , Ebran Nathalie , Gal Jocelyn , Otto Josiane , Giacchero Damien , Borchiellini Delphine , Guigay Joel , Peyrade Frederique , Milano Gerard , Saada Esma
Published: August 2018

Background: Hyperprogressive disease (HPD) has been described in advanced solid tumors patients treated with anti PD1 anti PDL1 monotherapy (Champiat, Clin Can Res 2016; Saada-Bouzid, Ann Oncol, 2017). Our aim was to explore whether host constitutional variations in the nucleotide sequence of genes involved in immune response might be associated with the occurrence of HPD in patients treated with anti PD-(L)1.

Methods : From April to August 2017, 98 patients were treated in the Centre Antoine Lacassagne (Nice, France) with anti PD-(L)1. Potential SNPs for four candidate genes with a minor allele frequency≥ 5% in caucasians were selected according to genome browser data-base: PD1 (rs10204525, rs11568821, rs2227981), PD-L1 (rs2282055, rs2297136, rs2297137, rs4143815, rs10815225), IDO1 (rs3739319, rs3808606, rs373931, rs9657182) and VEGR2 (rs2305948, rs1870377, rs2071559). High through-output genotyping was done by MassARRAY (AGENA). HPD was defined as a TGKR (ratio of the slope of tumor growth before treatment and the slope of tumor growth on treatment) ≥ 2.

Results: Patients were treated for head and neck squamous cell carcinoma (16/98), non-small cell lung cancer (48/98), melanoma (13/98), renal cell carcinoma (14/98) and 7 others. Median age was 66 years old, 33 were male. All tested SNPs were in Hardy-Weinberg equilibrium. Responses were associated with G3-G4 toxicities (10% vs 42%, p=0.001). TGKR was assessable in 80 patients. HPD was observed in 11/80 (14 %) patients. HPD was significantly associated with age≥70y (6% vs 25%, p=0.02) and genotype for PD1 rs 2227981 (8/32=35% G/G vs 2/43=5% A/G or A/A, p=0.02), PDL1 rs 2282055 (1/37=3% T/T vs 10/43=23% T/G or G/G,p=0.02) and VEGFR2 rs1870377 (2/48=5% T/T vs 9/32=28% T/A or A/A, p=0.005). Multivariate analysis confirmed the association between HPD and VEGFR2 rs1870377 T/A or A/A (OR=15.3, p=0.007), PDL1 rs 2282055 T/G or G/G (OR=18.7, p=0.01) and age≥70y (OR=14.4, p=0.006).). Combining the three alleles at risk for HPD gave an OR =12.4 (p=0.001).

Conclusion: In patients treated with anti PD-(L)1, HPD was independently associated with older age and host related gene variations. Host related immunogenetics could become an integrative part of predictive factor for immunotherapy outcome.

Citation Format: Sadal Refae, Nathalie Ebran, Jocelyn Gal, Josiane Otto, Damien Giacchero, Delphine Borchiellini, Joel Guigay, Frederique Peyrade, Gerard Milano, Esma Saada. Host immunogenetics and hyperprogression under PD1/PD-L1 checkpoint inhibitors [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 4548.
Abstract 4598: A comprehensive assessment of the impact of preanalytical variables on cell free DNA and circulating tumor cells in blood
Authors: Pottekat Anita , Allawi Hatim T. , Boragine Genna T. , Kaiser Michael W. , Sander Tamara , Krueger Chateen , Bruinsma Janelle J. , Murray Amber N.
Published: August 2018

Liquid biopsy continues to gain traction as a minimally invasive method to monitor biomarkers associated with malignancy and metastasis. A simple blood draw can reveal key biomarkers such as circulating tumor cells (CTCs) and cell-free DNA (cfDNA). However, these analytes are present in low quantities and are prone to rapid degradation and contamination, which presents challenges for the detection of rare CTCs and cfDNA targets. Higher molecular weight genomic DNA from poorly stabilized leukocytes can contaminate plasma, compromising liquid biopsy assay results by increasing the total amount of extracted DNA while diluting rare cfDNA targets. Preservation of CTCs and cfDNA over time is thus essential for the accuracy of downstream tests, and preanalytical variables that impact plasma volume, cfDNA yield and quality, and cell recovery in ex vivo blood must be understood. Here, we identified several preanalytical variables, including type of blood collection tube, storage time and conditions post-phlebotomy, and centrifugation conditions for plasma fractionation. We then carried out a series of studies to better understand the interplay between these preanalytical variables and plasma recovery, cell recovery, and cfDNA preservation in healthy donor blood, contrived samples, and patient samples. We found that choice of blood collection tube, in particular, has a significant impact on preservation of plasma volume, CTCs, cfDNA over time, particularly under real-world stress conditions including temperature excursions, sample shipping, and storage time post-draw. The work presented here demonstrates the importance of assessing preanalytical variables for the preservation of cancer biomarkers in whole blood without compromising sample integrity. This will ultimately help researchers make informed decisions on how best to stabilize their analytes of interest without introducing un-intended variables due to blood tube selection, shipping and storage conditions, and plasma isolation conditions.

Citation Format: Anita Pottekat, Hatim T. Allawi, Genna T. Boragine, Michael W. Kaiser, Tamara Sander, Chateen Krueger, Janelle J. Bruinsma, Amber N. Murray. A comprehensive assessment of the impact of preanalytical variables on cell free DNA and circulating tumor cells in blood [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 4598.
Abstract 462: Hyperactivation of ERK/MAPK pathway reduces pancreatic cancer cells' proliferation and tumor progression through phosphoproteome reprogramming
Authors: Rowell Marie-Camille , Deschênes-Simard Xavier , Calvé Benjamin Le , Lopes-Paciência Stéphane , Ruiz Ana Fernandez , Kottakis Filippos , Bardeesy Nabeel , Ferbeyre Gerardo
Published: August 2018

This study aims to better understand the loss of tumor -uppression functions of ERK in pancreatic cancer progression. Pancreatic cancer is the 4th leading cause of death by cancer in Canada and USA with a 5-year survival rate below 7%. We have identified a high level of phosphorylation of the MAP kinase ERK in benign pancreatic intraepithelial neoplasms (PanIN), followed by a reduction in phospho-ERK levels in pancreatic ductal adenocarcinoma (PDAC) both in human patient samples and KRas-driven mouse models for pancreatic cancer. Since PanINs are considered senescent lesions, we hypothesized that phospho-ERK levels were reduced in PDAC to bypass oncogene-induced senescence and allow tumor initiation. We propose that reactivating high ERK in PDAC could restore tumor-suppression mechanisms lost during progression. To do so, we use a tamoxifen-inducible constitutive allele of RAF1 kinase, which acts upstream of ERK (ΔRAF1-ERT). In human (PANC-1, KP-4) and murine PDAC cell lines, activated RAF1 slows cell proliferation and induces cellular senescence in vitro in an ERK-dependent and p53/p16-independent manner. Activation of ERK kinases by this system in subcutaneous PDAC xenografts slows tumor growth in nude mice. To characterize key mediators of ERK-dependent tumor suppression, we did phosphoproteomics by mass spectrometry and identified a reprogramming of nuclear vs. cytoplasmic phosphoproteome, involving key ribosome biogenesis factors. Depletion of these nucleolar targets by shRNAs and nucleolar stress inducers such as RNA polymerase I inhibitor CX-5461 recapitulates ERK-dependent tumor suppression. At the same time, we are also investigating autophagy-related processes involved in RAF/MEK signaling blockade in PDAC cells. In summary, we suggest that high levels of p-ERK1/2 in early PanIN stage act as a tumor-suppressor mechanism, which is lost with cancer progression. Our future work aims to better understand the mechanism by which high ERK signaling exerts tumor suppression in order to find downstream effectors that could be used as therapeutic targets in pancreatic cancer.

Citation Format: Marie-Camille Rowell, Xavier Deschênes-Simard, Benjamin Le Calvé, Stéphane Lopes-Paciência, Ana Fernandez Ruiz, Filippos Kottakis, Nabeel Bardeesy, Gerardo Ferbeyre. Hyperactivation of ERK/MAPK pathway reduces pancreatic cancer cells' proliferation and tumor progression through phosphoproteome reprogramming [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 462.
Abstract 4637: Tumor treating fields (TTFields) have antiproliferative effects on high-grade pediatric brain tumor cell lines
Authors: Branter Joshua , Estevez-Cebrero Maria , Grundy Richard , Basu Surajit , Smith Stuart
Published: August 2018

Introduction Approximately 400 children in the UK are diagnosed with a CNS tumor each year, which represents a quarter of childhood cancer cases in the UK. The 5-year survival rate of paediatric brain tumor patients is approximately 75%, but survival remains much worse for some brain tumor patients, particularly high-grade gliomas. Pre-clinical and clinical studies have demonstrated efficacy of TTFields as a treatment for high-grade glioma leading to the approval of TTFields for adult Glioblastoma multiforme (GBM) patients. The lack of overlapping toxicities associated with TTFields has made electric field therapy an attractive treatment strategy for poor prognosis paediatric brain tumors.Methods The inovitro is the laboratory based testing system used to deliver TTFields to cell cultures. inovitro was used to deliver TTFields over 72 hours at a range of clinically relevant frequencies (100-400kHz) to our panel of paediatric GBM, Medulloblastoma and Ependymoma cell lines. The effects of TTFields on cell viability and cell cycle was assessed using Resazurin based metabolic viability tests and flow cytometry. The determined optimal frequency for each cell line was also combined with a mitotic inhibitor to explore synergistic effects. Gene expression changes of TTFields treated cells was investigated. Results TTFields treatment reduces cell viability of all cell lines tested within the frequency panel. The optimum frequencies were between 100 and 400kHz for all cell lines tested. TTFields treatment had effects on cell cycling with increased G2 accumulation of treated cells being observed. Combining TTFields with mitotic inhibitors increased efficacy in an additive fashion. Conclusions This preliminary study has demonstrated efficacy of TTFields on paediatric brain tumor cell lines. The optimal frequencies for all the paediatric GBM cell lines used did not match the 200kHz as previously reported for U-87 MG cells. This report warrants further study into the potential for TTFields therapy for paediatric brain tumor patients.

Citation Format: Joshua Branter, Maria Estevez-Cebrero, Richard Grundy, Surajit Basu, Stuart Smith. Tumor treating fields (TTFields) have antiproliferative effects on high-grade pediatric brain tumor cell lines [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 4637.
Abstract 4642: Personalized cancer-specific protein corona affects the therapeutic impact of nanoparticles
Authors: Corbo Claudia , Mahmoudi Morteza , Farokhzad Omid C.
Published: August 2018

Despite recent progress in nanomedicine, there is still a lack of efficient nanotherapeutics for cancer treatment. When nanoparticles (NPs) are injected, they circulate the bloodstream and interact with biomolecules, creating a biomolecular shell. This shell of biomolecules surrounding the NPs consists mostly of proteins and is referred to as the protein corona (PC). The PC composition depends on various factors, including, but not limited to i) surface properties of NPs (e.g. surface chemistry and charge), and ii) experimental parameters (e.g. incubation time, pH, temperature). The impact of these factors on the formation of the PC has been widely investigated and is nowadays well known. However, the effects of the biological milieu (e.g. patient's health status, plasma vs serum) on the PC formation and composition have been underexamined and still need to be deeply clarified. Recently, it has been demonstrated that the protein pattern constituting the PC of NPs exposed to plasma of cancer patients is different than that of the PC formed by exposure of NPs to healthy plasma. These variations can be miniscule but crucial in the fate of NPs. In fact, NPs surrounded by a PC lose their synthetic identity and acquire a new biological identity responsible for their biological destiny (cell targeting, accumulation, immune response). In this work, we have studied the PCs formed around NPs using plasmas of patients with one of eight cancers to gain insights into the cancer-related variations of the PC composition and into their potential effects on the therapeutic efficacy of NPs. Our results confirmed that the same NPs incubated with plasmas of patients affected by different tumors have distinct PCs. Overall, this is the first wide study unveiling the specific PCs formed around NPs using plasmas of eight cancers. This comprehensive report acts as a tool for researchers in nanomedicine to design personalized nanotherapeutics in a cancer-specific manner for clinical applications. Methods: Silica NPs were incubated with plasma of healthy subjects and cancer patients. NPs in the study were 100 nm in size and the PCs in each subject's plasma were characterized and compared using SDS-PAGE and LC-MS/MS. Plasmas of patients with one of 8 cancers have been employed: breast, rectum, lung, kidney, thyroid, uterine, bladder, ovary.

Citation Format: Claudia Corbo, Morteza Mahmoudi, Omid C. Farokhzad. Personalized cancer-specific protein corona affects the therapeutic impact of nanoparticles [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 4642.
Abstract 4713: BET bromodomain inhibition synergizes with PD-1 blockade to facilitate anti-tumor response in Kras-mutant non-small cell lung cancer
Authors: Adeegbe Dennis O. , Liu Shengwu , Bowden Michaela , Hammerman Peter S. , Bradner James E. , Rustgi Anil K. , Bass Adam J. , Freeman Gordon J. , Chen Huawei , Wong kwok-Kin
Published: August 2018

KRAS mutation is present in about 30% of human lung adenocarcinomas. While recent advances in targeted therapy have shown great promise, KRAS remains undruggable and concurrent alterations in tumor suppressors render KRAS mutant tumors even more resistant to existing therapies. Contributing to the refractoriness of KRAS mutant tumors harboring these co-mutations are immunosuppressive mechanisms such as increased presence of suppressive Tregs in tumors and elevated expression of the inhibitory receptor PD-1 on tumor-infiltrating T cells. BET bromodomain inhibitors demonstrate clinical benefit in hematologic malignancies, and prior reports demonstrate their Treg-disruptive effects in a NSCLC model. Targeting PD-1 inhibitory signals through anti-PD-1 antibody blockade has also shown substantial therapeutic impact in lung cancer although these outcomes are still limited to a minor pool of patients. We therefore hypothesized that the BET bromodomain inhibitor JQ1 would synergize with PD-1 blockade to promote robust anti-tumor response in lung cancer. In the present study, using Kras+/LSL-G12D; Trp53L/L (KP) mouse models of non-small cell lung cancer, we identified cooperative effects among JQ1 and anti-PD-1 that included reduced numbers of tumor-infiltrated Tregs and enhanced activation of tumor-infiltrating T cells, which exhibited a Th1 cytokine profile that favored their demonstrated improved effector function. Furthermore, lung-tumor-bearing mice under this combinatorial treatment regimen showed robust and long-lasting anti-tumor responses compared to either agent alone, culminating in substantial improvement in the survival of treated mice. Thus, combining BET bromodomain inhibition with immune checkpoint blockade offers a promising therapeutic approach for solid malignancies such as lung adenocarcinoma.

Citation Format: Dennis O. Adeegbe, Shengwu Liu, Michaela Bowden, Peter S. Hammerman, James E. Bradner, Anil K. Rustgi, Adam J. Bass, Gordon J. Freeman, Huawei Chen, kwok-Kin Wong. BET bromodomain inhibition synergizes with PD-1 blockade to facilitate anti-tumor response in Kras-mutant non-small cell 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 4713.