bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4867: Treating hepatocellular carcinoma metastasis and overcoming chemoresistance through inhibiting the MDM2 oncogene
Authors: Wang Wei , Hu Bo , Qin Jiang-Jiang , Cheng Jianwen , Li Xin , Hu Ming , Buolamwini John , Yang Xin-Rong , Fan Jia , Zhang Ruiwen
Published: August 2018

Hepatocellular carcinoma (HCC) poses a major health problem worldwide and novel, effective therapeutic approaches are urgently needed. The MDM2 oncogene is amplified and overexpressed in HCC and associated with its initiation, progression, metastasis, and chemoresistence. MDM2 has been demonstrated to be as a promising therapeutic target for the treatment of HCC. However, most of the current MDM2 inhibitors have been designed to block the binding between MDM2 and p53, and have limited efficacy against tumors with mutant or deficient p53. We recently performed a high-throughput virtual and cell-based screening, yielding a novel MDM2 inhibitor, termed SP141. The present study was designed to evaluate the anti-HCC activity of SP141, its potential to chemosensitize HCC cells to other anticancer agents, and to determine the underlying mechanism(s) of action of SP141 in various in vitro and in vivo models with different backgrounds of p53 (wildtype, mutant or null). Our results demonstrated that SP141 inhibited cell growth and prevented cell migration and invasion, independent of p53. Mechanistically, SP141 directly bound to the MDM2 protein and promoted MDM2 degradation. The inhibition of MDM2 by SP141 also increased the sensitivity of HCC cells to sorafenib. In addition, in orthotopic and patient-derived xenograft (PDX) models, SP141 inhibited MDM2 expression and suppressed tumor growth and metastasis, without any host toxicity. Furthermore, mechanistic study demonstrated that the inhibition of MDM2 by SP141 was essential for its anti-HCC activities. In conclusion, MDM2 inhibition by SP141 resulted in the inhibition of HCC tumor growth and metastasis and sensitization of HCC cells to chemotherapy, regardless of their p53 status. These results provide support for the further development of SP141 as a lead candidate for the treatment of HCC. (Supported by NIH R01 CA186662 and R01CA214019 and ACS RSG-15-009-01-CDD.)

Citation Format: Wei Wang, Bo Hu, Jiang-Jiang Qin, Jianwen Cheng, Xin Li, Ming Hu, John Buolamwini, Xin-Rong Yang, Jia Fan, Ruiwen Zhang. Treating hepatocellular carcinoma metastasis and overcoming chemoresistance through inhibiting the MDM2 oncogene [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 4867.
Abstract 4873: Therapeutic targeting of LSD1 in Ewing sarcoma with SP-2509
Authors: Pishas Kathleen , Boone Megann , Sharma Sunil , Lessnick Stephen
Published: August 2018

Objective: Multimodal regimes remain the cornerstone treatment for Ewing sarcoma (ES), the second most common solid bone malignancy diagnosed in pediatric and young adolescent populations. We have reached a therapeutic ceiling with conventional cytotoxic agents, highlighting the need to adopt novel approaches specifically targeting the drivers of ES oncogenesis. As LSD1 (Lysine Specific Demethylase 1) is highly expressed in ES tumors, with elevated expression levels associated with worse overall survival, this study examined biomarkers of sensitivity/mechanisms of cytotoxicity to targeted LSD1 inhibition using SP-2509 (reversible LSD1 inhibitor).

Methods: The antiproliferative effects of SP-2509 were determined through Cell Titre Glo assays following 72hrs of treatment in a panel of 17 ES cell lines with varying STAG2/TP53 mutational status and basal LSD1 expression levels. RNA-seq analysis of six ES cell lines -/+ SP-2509 treatment (2μM) was also conducted.

Results: Innate resistance to SP-2509 was not observed in our cell line cohort (72hr IC50:81nM-1593nM). In contrast, resistance to the clinical next-generation LSD1 irreversible inhibitor GSK-LSD1 was observed across multiple cell lines (144hr IC50>300μM). Although TP53/STAG2 mutational status and basal LSD1 mRNA/protein levels did not correlate with SP-2509 response, induction of LSD2 (homologue of LSD1) following SP-2509 treatment was strongly associated with SP-2509 hypersensitivity (R2=0.562). shRNA mediated knockdown of LSD2 significantly reduced the cytotoxic effects of SP-2509 (4.3 fold IC50 increase) only in hypersensitive cell lines. Mechanistically, RNA-seq analysis revealed that SP-2509 imparts apoptosis through engagement of the endoplasmic reticulum stress pathway, with hypersensitive cell lines sharing similar transcriptomic profiles. In addition, ETS1/HIST1H2BM were specifically induced/repressed, respectively, following SP-2509 treatment only in our hypersensitive cell lines. Finally, we demonstrate that the transcriptional profile driven by SP-2509 strongly mirrors LSD1 genetic depletion.

Conclusion: Our findings provide key insights into the mechanisms of SP-2509 cytotoxicity as well as biomarkers that can be used to predict LSD1 inhibitor sensitivity in ES.

Citation Format: Kathleen Pishas, Megann Boone, Sunil Sharma, Stephen Lessnick. Therapeutic targeting of LSD1 in Ewing sarcoma with SP-2509 [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 4873.
Abstract 489: Up-regulation of miR-130b is involved in prostate cancer racial disparity through FHIT pathway inactivation
Authors: Hashimoto Yutaka , Shiina Masrisa , Kato Taku , Yamamura Soichiro , Tanaka Yuichiro , Majid Shahana , Saini Sharanjot , Shahryari Varahram , Kulkarni Priyanka , Dasgupta Pritha , Bhagiratha Divya , Bhat Nadeem , Deng Guoren , Tabatabai Laura , Kumar Deepak , Dahiya Rajvir
Published: August 2018

Purpose: Prostate cancer (PCa) is a common cancer in men. African-American (AfA) men have higher incidence and significantly higher prostate cancer mortality rates than Caucasian-American (CaA) men. In this study, we investigated the biochemical role of miR-130b in this disparity.

Methods: We used meta-analyses and data from Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA). PCa cell lines from CaA (DU145, LNCaP, PC3), AfA (MDA-PCa-2b, E006AA-hT) and a normal epithelial cell line (PWR1E) were used for examining miR-130b expression levels and/or establishing stable miR-130b knock-down clones. Total RNA was extracted from clinical samples and cell lines. miR-130b and mRNA levels of its target genes were determined by quantitative real-time PCR (qPCR). qPCR based gene expression arrays were used to identify miR-130b target genes. Cell proliferation, apoptosis and cell cycle were monitored after stable miR-130b knock down. Western blotting was performed to detect miR-130b target protein expression levels.

Results: TCGA data showed up-regulation of miR-130b in AfA PCa tissue samples compared with CaA samples. miR-130b was significantly up-regulated in AfA prostate cancer tissues and cell lines compared to CaA cells and tissues. Utilizing SFVAMC and NDRI patient cohorts, we confirmed that miR-130b expression was linked to a racial difference between AfA/CaA PCa patients. Knock down of miR-130b showed decreased growth and cell cycle arrest, though the effect was less in CaA compared to AfA cells. We found unique changes in biological pathways associated with miR-130b knock down in AfA cells by qPCR array. Evaluation of the altered pathways showed that Fragile Histidine Triad (FHIT) was markedly changed in the AfA compared with CaA cells.

Conclusion: These results demonstrate that miR-130b may be a central regulator of key events that contribute racial differences in prostate cancer. FHIT may also be a new AfA specific tumor-suppressive pathway that may be of therapeutic importance in AfA prostate cancer patients.

Citation Format: Yutaka Hashimoto, Masrisa Shiina, Taku Kato, Soichiro Yamamura, Yuichiro Tanaka, Shahana Majid, Sharanjot Saini, Varahram Shahryari, Priyanka Kulkarni, Pritha Dasgupta, Divya Bhagiratha, Nadeem Bhat, Guoren Deng, Laura Tabatabai, Deepak Kumar, Rajvir Dahiya. Up-regulation of miR-130b is involved in prostate cancer racial disparity through FHIT pathway inactivation [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 489.
Abstract 4910: Development of an antagonist to the TWEAK-Fn14 ligand-receptor interaction with enhanced efficacy/potency in glioblastoma cells
Authors: Harder Bryan , Sereduk Chris , Dhruv Harshil , Millard Nghia , Kim Anthony , Winkles Jeffrey , Woodworth Graeme , Yin Holly , Tran Nhan L.
Published: August 2018

Glioblastoma multiforme (GBM) remains the most common and deadly primary brain tumor with minimal advances in therapeutic development for pharmacological treatment. Tumor resection followed by temozolomide (TMZ) and radiotherapy adds months to survival; however, tumor recurrence usually occurs. The fibroblast growth factor-inducible 14 (Fn14) receptor has been shown to be up-regulated in the GBM cells left behind after surgery and contributes to an invasive and pro-survival phenotype, providing a desirable molecule to target after removal of the primary tumor. Previously, L524-0366 was reported by our lab to be the first antagonist of Fn14 signaling; however, minimal efficacy/potency has limited this drug from progressing from a useful research tool to preclinical studies. Structure activity relationship (SAR) studies using the parent L524-0366 compound gave rise to K78, which was found to bind to Fn14 with enhanced affinity, while showing no binding to the structurally-related TNFα receptor. Furthermore, K78 was able to abate the downstream signaling effects initiated by the Fn14 ligand, tumor necrosis factor-like weak inducer of apoptosis (TWEAK), as well as sensitize GBM cells to the cytotoxic effects of TMZ. TWEAK-induced cellular migration was also mitigated with co-treatment of K78 in GBM cells, compared to a vehicle control. It is anticipated that further development of Fn14 antagonists will be of great importance for the advancement of GBM patient treatment and warrant further investigation as primary and combination therapy, especially for the prevention of recurrent disease.

Citation Format: Bryan Harder, Chris Sereduk, Harshil Dhruv, Nghia Millard, Anthony Kim, Jeffrey Winkles, Graeme Woodworth, Holly Yin, Nhan L. Tran. Development of an antagonist to the TWEAK-Fn14 ligand-receptor interaction with enhanced efficacy/potency in glioblastoma 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 4910.
Abstract 4926: Preclinical intratumoral pharmacokinetics (PK) of capecitabine (Cap) given +/- eribulin (Eri)
Authors: Casulleras Maria Jove , Spencer Jade , Loadman Paul , Clench Malcolm , Shnyder Steve , Cooper Patricia , Russo Cristina , Race Amanda , Salazar Ramon , Twelves Chris
Published: August 2018

Background: Eri & Cap are active as single agents in metastatic breast cancer, but the combination appears particularly active & well tolerated clinically. We explored the hypothesis that normalisation of tumor vasculature by Eri seen in preclinical models may enhance delivery of Cap to the tumor when given in combination.

Methods: Mice bearing MDA-MD-231 xenografts were treated with Cap 540mg/kg p.o. day 1 alone or following Eri 1mg/kg i.v. on day -8 then sacrificed on day 1 at 15min, 30min, 1h, 2h, 4h, 6h or 24h after Cap (n=3/ time point). Concentrations of Cap & its metabolites (5'-deoxy-5-fluorocytidine, 5DFCR; 5-Fluoro-5'-deoxyuridine, 5DFUR; 5-Fluorouracil, 5FU; 5-Fluorouridine, 5FUrd; & 5-Fluoro-2'-deoxyuridine, 5FdUrd) in plasma, tumor, skin & muscle were analysed by LC-MS/MS. Half the tumor was reserved to study Cap distribution using matrix-assisted laser desorption/ionization mass spectrometry imaging with haem imaged as a surrogate marker for tumour vasculature.

Results: We found no differences in the plasma, skin, muscle or tumor PK profiles of Cape & its metabolites with the addition of Eri (Table 1). In both treatment arms intratumoral concentrations of 5DFUR, 5FU, 5FUrd & 5FdUrd appeared higher in tumors while those of Cape & 5DFCR appeared higher in normal healthy tissues. To mitigate the variability in tumor Cape concentrations between animals, we also expressed the results as the ratio of drug concentration in tumour:healthy tissue but again found no apparent effect of Eri on intratumoral drug concentrations. Analyses of drug distribution are on-going & will be presented.

Conclusion: No clear effect of Eri on intratumoral concentrations of Cape or its metabolites was seen. We are exploring potential differences in tumour drug distribution.

Citation Format: Maria Jove Casulleras, Jade Spencer, Paul Loadman, Malcolm Clench, Steve Shnyder, Patricia Cooper, Cristina Russo, Amanda Race, Ramon Salazar, Chris Twelves. Preclinical intratumoral pharmacokinetics (PK) of capecitabine (Cap) given +/- eribulin (Eri) [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 4926.
Abstract 4935: High-throughput immune-oncology screen identifies EGFR inhibitors as potent enhancers of CTL antigen-specific tumor cell killing
Authors: Lizotte Patrick H. , Luster Troy , Cavanaugh Megan E. , Taus Luke J. , Dhaneshwar Abha , Mayman Naomi , Yang Aaron , Bittinger Mark , Kirschmeier Paul , Gray Nathanael S. , Barbie David A. , Janne Pasi A.
Published: August 2018

As immune checkpoint blocking antibodies increasing become foundational therapies for the treatment of cancer, there is a pressing need to identify compounds that synergize with checkpoint blockade as the basis of combinatorial treatment regimens. We have developed a screening assay in which a luciferized tumor cell line expressing a model antigen is co-cultured with a transgenic CD8+ T cell specifically recognizing the model antigen in a H-2b-restricted manner. The target tumor cell/T cell assay was screened with a small molecule library to identify compounds that inhibit or enhance T cell-mediated killing of tumor cells in an antigen-dependent manner. The EGFR inhibitor Erlotinib was the top hit that enhanced T cell killing of tumor cells. Subsequent experiments with Erlotinib and additional EGFR inhibitors validated the screen result. EGFR inhibitors increase both basal and IFN-γ-induced antigen processing and presentation of MHC class-I, which enhanced recognition and lysis by CD8+ cytotoxic T lymphocytes. The tumor cell line was also transduced to constitutively express Cas9, and a pooled CRISPR screen utilizing the same target tumor cell/T cell assay identified sgRNAs targeting EGFR as sensitizing tumor cells to T cell-mediated killing. Combination of PD-1 blockade with EGFR inhibition showed significant synergistic efficacy in the MC38 syngeneic colon cancer model that was superior to PD-1 blockade or EGFR inhibition alone, further validating EGFR inhibitors as immunomodulatory agents that enhance PD-1 checkpoint blockade. This novel target tumor cell/T cell assay can be screened in high-throughput with small molecule libraries and genome-wide CRISPR/Cas9 libraries to identify both compounds AND target genes, respectively, that enhance or inhibit T cell recognition and killing of tumor cells.

Citation Format: Patrick H. Lizotte, Troy Luster, Megan E. Cavanaugh, Luke J. Taus, Abha Dhaneshwar, Naomi Mayman, Aaron Yang, Mark Bittinger, Paul Kirschmeier, Nathanael S. Gray, David A. Barbie, Pasi A. Janne. High-throughput immune-oncology screen identifies EGFR inhibitors as potent enhancers of CTL antigen-specific tumor cell killing [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 4935.
Abstract 4982: Oncogenic role of THOR, a conserved cancer/testis long noncoding RNA
Authors: Niknafs Yashar , Hosono Yasuyuki , K Matthew , Prensner John , Mehra Rohit , Pitchiaya Sethuramasundaram , Tien Jean , Malik Rohit , Zhao Weibin , Chinnaiyan Arul
Published: August 2018

Large scale transcriptome sequencing efforts have vastly expanded the catalog of long non-coding RNAs (lncRNAs) with varying evolutionary conservation, lineage expression, and cancer specificity. Through a recent large-scale transcriptomic analysis over 6,500 tumor RNA-seq samples, we discovered over 50,000 lncRNAs in the human genome, many of which exhibited highly evolutionarily conserved sequence patterns. Building upon the discovery of these highly conserved lncRNAs, we functionally characterized a novel ultraconserved lncRNA, THOR, which exhibits expression exclusively in testis and a broad range of human cancers. We establish THOR as the first discovered cancer/testis lncRNA, and further investigated its functional significance. THOR knockdown and overexpression in multiple cell lines and animal models alters cell or tumor growth supporting an oncogenic role. Namely, we generated CRISPR knockout cell line models, showing a definitive role for THOR in cancer progression. Additionally, given the sequence conservation of THOR through the mouse and zebrafish, we generated a zebrafish knockout model, and also a zebrafish overexpression model for THOR. Through RNA-pulldown followed by mass spectrometry, we discovered a conserved interaction of THOR with the RNA binding protein, IGF2BP1, in both human and zebrafish cells. We further show that THOR contributes to the mRNA stabilization activities of IGF2BP1. These findings are corroborated by iCLIP data for IGF2BP1. Notably, transgenic THOR knockout produced fertilization defects in zebrafish and also conferred a resistance to melanoma onset in an NRAS K61-induced zebrafish melanoma model. Likewise, ectopic expression of human THOR in zebrafish accelerated the onset of melanoma. THOR represents a novel class of functionally important cancer/testis lncRNAs whose structure and function have undergone positive evolutionary selection.

Citation Format: Yashar Niknafs, Yasuyuki Hosono, Matthew K, John Prensner, Rohit Mehra, Sethuramasundaram Pitchiaya, Jean Tien, Rohit Malik, Weibin Zhao, Arul Chinnaiyan. Oncogenic role of THOR, a conserved cancer/testis long noncoding RNA [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 4982.
Abstract 499: MicroRNA-411 downregulation promotes tumorigenesis and inhibits chemotherapy sensitivity by upregulating MLLT11 expression in human bladder cancer
Authors: Jin Honglei , Huang Haishan , Huang Chuanshu , Hua Xiaohui , Zhu Junlan
Published: August 2018

Although several previous studies have reported the implication of various microRNAs (miRNAs) in regulation of human bladder cancer (BC) development, alterations and function of many miRNAs in BC carcinogenesis and anti-cancer therapy are not explored yet at present. Here, we screened 1900 known miRNAs and first discovered that miR-411 was one of major miRNAs that was downregulated in a n-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced BC mouse model. This miR-411 downregulation was also observed in human BC tissues and cell lines. The results from evaluating the relationship between miR-411 and patient survival in BC using the TCGA database also indicated that miR-411 was positively correlated with survival (>3 years). Our studies also showed that miR-411 inhibited tumorigenesis and increased chemotherapy sensitivity in human BC. Mechanistic studies revealed that overexpression of miR-411 repressed expression of ALL1-fused gene from the chromosome 1q (MLLT11) by binding to the 3′-untranslated region of MLLT11 mRNA, induced p21 expression, and caused cell cycle arrest at the G2/M phase, in turn inhibiting BC tumorigenic growth. Ectopic expression of miR-411 also increased the sensitivity of BC cells to the chemotherapeutic agents, whereas MLLT11 restoration in miR-411-overexpressing cells increased resistance to the chemotherapeutic agents.

Citation Format: Honglei Jin, Haishan Huang, Chuanshu Huang, Xiaohui Hua, Junlan Zhu. MicroRNA-411 downregulation promotes tumorigenesis and inhibits chemotherapy sensitivity by upregulating MLLT11 expression in human bladder 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 499.
Abstract 5005: Aging promotes changes to peritoneal and omental collagen structure that contribute to increased ovarian cancer metastatic success
Authors: Harper Elizabeth , Loughran Elizabeth , Leonard Annemarie , Hilliard Tyvette , Asem Marwa , Liu Yueying , Yang Jing , Klymenko Yuliya , Johnson Jeff , Sheedy Emma , Shi Zonggao , Leevy Matthew , Ravosa Matthew , Stack M. Sharon
Published: August 2018

Ovarian cancer (OvCa) is the deadliest cancer of the female reproductive system, ranking fifth in overall cancer deaths among women. Unlike the majority of cancers, OvCa metastasizes via diffusion through the peritoneal cavity, resulting in multiple metastatic sites, including the omentum and peritoneum. These metastasizing OvCa cells induce rapid mesothelial cell retraction and readily adhere to the sub-mesothelial collagen of the extracellular matrix. Epidemiologic data identifies age as a significant risk factor in OvCa, as about half of diagnoses are in women over the age of 63. Despite this, age is understudied in the OvCa field. Using a C57Bl/6 mouse model of aging, young (Y) mice ranging from 3-6 months of age, and aged (A) mice ranging from 20-23 months of age, corresponding to women aged 20-30 years (Y) and 60-67 years (A) were used to study the role aging has on metastasis. Fluorescently tagged C57Bl/6 syngeneic ID8 p53-/- mouse OvCa surface epithelial cells were injected intraperitoneally in young and aged mice and disease progression was evaluated for 5.5 weeks. Organ-specific tumor burden was quantified with ImageJ, revealing increased tumor burden in aged mice compared to their young counterparts. These results were reproduced in the FVB mouse model using syngeneic PTENshRNA/KRASG12V modified FVB OvCa oviductal epithelial cells. Second Harmonic Generation Microscopy (SHG) was used to visualize collagen of the peritoneal and omental tissues from young and aged C57Bl/6 mice. Distinct structural differences were shown in omental collagen in the Y vs A cohorts and validated with Scanning Electron Microscopy (SEM). Additionally, Nanoindentation illustrated mechanical differences between young and aged peritoneal samples. In conclusion, aging induces changes in the structure and mechanical strength of peritoneal and omental collagen, which contribute to OvCa metastasis.

Citation Format: Elizabeth Harper, Elizabeth Loughran, Annemarie Leonard, Tyvette Hilliard, Marwa Asem, Yueying Liu, Jing Yang, Yuliya Klymenko, Jeff Johnson, Emma Sheedy, Zonggao Shi, Matthew Leevy, Matthew Ravosa, M. Sharon Stack. Aging promotes changes to peritoneal and omental collagen structure that contribute to increased ovarian cancer metastatic success [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 5005.
Abstract 5024: Drug screening of biopsy-derived multicellular spheroids using microfluidic technology
Authors: Mulholland Theresa E. , McAllister Milly , Patek Samantha , Flint David , Underwood Mark , Sim Alexander , Edwards Joanne , Zagnoni Michele
Published: August 2018

Performing drug screening, of physiologically relevant three-dimensional (3D) tumor models, for personalized treatment remains challenging, due to the small amount of tissue available from most biopsies. New microfluidic technologies, enabling greater control over cell positioning and fluid behavior at the micro-scale, allow extensive testing of anticancer agents on human tumor tissue preparations in 3D and offer new solutions for the development of anticancer compounds and personalized medicine. We have developed a microfluidic platform for extensive drug screening of tumor biopsies in a cost-effective manner and validated the system with tumor prostate patient samples. As a typical drug screening assay, up to 22 drug concentration-response curves could be generated from a single biopsy, within a time frame of up to 4 weeks. Biopsy tissue, grown as a heterogeneous co-culture from the primary sample, was prepared as cancer-cell enriched multicellular spheroids, cultured for 3 to 5 days prior to the application of a panel of standard-of-care drugs for prostate cancer. Readouts were obtained via bright-field and epifluorescence microscopy. The microfluidic platform was designed to be operated entirely without the need of external fluid actuation, with the microfluidic network capable of generating long-lasting, stable and repeatable drug concentration gradients across arrays of 240 spheroids. Outcomes were generated as 8-point drug concentration response curves per device, with each drug concentration tested on at least 24 spheroids. In-house developed software was used to analyze bright-field and fluorescent images to provide readouts of spheroid growth and viability, as well as information on drug penetration and drug efficacy over time. Following platform and assay validation using cancer cell lines, proof-of-concept screening was performed on prostate biopsies from 2 different patients. Results showed that biopsy-derived spheroids were more resistant to treatment than LNCaP spheroids, a prostate cancer cell line. For one biopsy, spheroids were sensitive to docetaxel, but resistant to enzalutamide, despite the presence of intact androgen receptors. This preliminary data outlines how this technology could become a useful tool to investigate patient-specific drug resistance and to test novel anticancer agents in a cost-effective manner, based on maximized screening of human tumor tissue in a 3D format.

Citation Format: Theresa E. Mulholland, Milly McAllister, Samantha Patek, David Flint, Mark Underwood, Alexander Sim, Joanne Edwards, Michele Zagnoni. Drug screening of biopsy-derived multicellular spheroids using microfluidic technology [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 5024.
Abstract 5032: Tumor in a Dish (TiD): Novel approach for precision therapy using patient-derived cells
Authors: Ramamoorthy Prabhu , Thomas Sufi , Ramalingam Sathish , Fnu Gaurav , Sittampalam Sita G. , Jensen Roy A. , Anant Shrikant
Published: August 2018

Metastatic colorectal cancer is a major cause of cancer-related deaths. Most patients with metastatic disease fail to respond to current chemotherapeutic agents. Current, traditional culture model is not appropriate to study the metastatic disease in in vitro condition. Hence, there is an ominous need for developing new therapeutic and preventive agents that can target metastatic cancer. Unfortunately, there are no good in vitro as well as in vivo models available for studying metastatic disease. Hence, in this abstract, we demonstrate the development of a new lung model system, called Primitive Lung in a Dish or PLiD with cancer cells called “Tumor in a Dish” or TiD. The PLiD system shows expression of specific cell type marker like E-cadherin (epithelial cell) vimentin (fibroblast), CD31 (heme-endothelial cells) and LYVE1 (lymph-endothelial cells). Interestingly, we showed expression of lung functional protein like surfactant protein B and D. In the multicell type TiD system, cancer cells were grown in a 3-dimensional (3D) PLiD system containing normal epithelial cells, fibroblasts and endothelial cells. The model resembles in vivo tumor microenvironments, including cell-cell contact, tumor architecture, and the influence of different cell types. We next determined the efficacy of standard colon cancer chemotherapeutic agents. We observed differential activity of 5FU in TiD system when compared to standard 2D and single-cell type 3D cultures. Another surprising result we found was with freshly isolated colon cancer cells from patient samples. Without knowing the genetic characteristics of the cancer tissue, the TiD system was able to identify cells that were resistant to oxaliplatin. Moreover, using this method, we have developed novel drugs that target cancer cells while not affecting the normal tissue. More importantly, the technique has significant utility in precision/personalized medicine, wherein this phenotypic screen can be coupled with current DNA pharmacogenetics to identify the ideal therapeutic agent, thereby increasing the probability of response to treatment while reducing unnecessary side effects.

Citation Format: Prabhu Ramamoorthy, Sufi Thomas, Sathish Ramalingam, Gaurav Fnu, Sita G. Sittampalam, Roy A. Jensen, Shrikant Anant. Tumor in a Dish (TiD): Novel approach for precision therapy using patient-derived 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 5032.
Abstract 5051: STAT3 signaling activates MSK1-mediated histone H3 phosphorylation in N-nitrosocompounds induced carcinogenesis
Authors: Qi Hongyan , Yang Zhiyi , Zhang Shuilian , Ke Xinxin , Dai Chujun , Shao Jimin , Shen Jing
Published: August 2018

Signal transducer and activator of transcription (STATs) signaling, particularly STAT3, have been demonstrated to be one of the central pathways for cancer initiation and progression. However, the functional role of STAT3 in regulation of epigenetic changes in cancer, such as histone modifications, remains unclear. Our present study performed on N-nitroso compounds (NOC) induced gastric epithelial cell transformation revealed a positive correlation of active STAT3 signaling with increased histone H3 phosphorylation. Further analysis showed that mitogen- and-stress activated protein kinase (MSK1) was upregulated by NOC treatment, thereby promoting H3 Ser 10 phosphorylation (H3S10P). Intriguingly, putative STAT3 binding sites were identified in the MSK1 promoter, and we finally confirmed that MSK1 is a novel target gene of STAT3. It has been reported that STAT3 is a direct substrate for MSK1. Therefore, we analyzed the status of STAT3 activation after MSK1 overexpression and knockdown, and a positive feedback loop was found between them. With specific siRNA or inhibitors, we further demonstrated the aberrant activation of STAT3/MSK1/H3S10P axis in MNU-induced gastric cancer (GC) model in mice, human gastric cancer cells, and GC patients. ChIP-seq analysis was performed to determine the H3S10 phosphorylation distribution pattern in NOC-transformed gastric epithelial cells, and some candidate regulated genes were successfully identified. Together, the results of these studies shed light onto a critical role of STAT3 signaling in epigenetic modification network, mediated at least in part by MSK1. Our results suggest that STAT3/MSK1/H3S10P pathway and downstream key effectors represent interesting therapeutic targets in gastric carcinogenesis.

Citation Format: Hongyan Qi, Zhiyi Yang, Shuilian Zhang, Xinxin Ke, Chujun Dai, Jimin Shao, Jing Shen. STAT3 signaling activates MSK1-mediated histone H3 phosphorylation in N-nitrosocompounds induced carcinogenesis [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 5051.
Abstract 5087: Human melanoma exosomes induce metabolic reprogramming in human adult dermal fibroblasts
Authors: Shu Shin La , Allen Cheryl L. , Yang Yunchen , Maguire Orla , Minderman Hans , Sen Arindam , Ciesielski Michael J. , Collins Katherine A. , Bush Peter J. , Morgan Martin , Wu Yun , Bankert Richard B. , Ernstoff Marc S.
Published: August 2018

The “seed and soil” hypothesis of cancer metastasis has led us to examine the role of tumor-derived exosomes on reprogramming stromal cells composing the tumor microenvironment. We used human adult dermal fibroblasts (HADF) in vitro as a model of the premetastatic niche. Specifically, we determined the role of human melanoma-derived exosomes (HMEX) on governing metabolic reprogramming of healthy HADF. HMEX from six melanoma cell lines (3 BRAF V600E mutant cell lines (526-mel, 888-mel and Hs 294T) and 3 BRAF wild-type cell lines (1300-mel, 2183-Her4 and HMCB)) were purified by size-exclusion chromatography. Using the fluorescent tag CellVue (Maroon) we demonstrated via ImageStream that HMEXs are rapidly taken up by HADF. Following 18 hours of exposure to HMEX, HADF increased aerobic glycolysis and reduced oxidative phosphorylation (OXPHOS) in vitro, as measured by Agilent Seahorse XFe96. We have also developed an immuno-biochip that captures exosomes via anti-CD63 antibodies and simultaneously detects exosomal microRNAs by cationic lipoplex nanoparticles (tCLN) containing molecular beacons. With this immuno-biochip, we demonstrated the presence of microRNAs miR-155 and miR-210 in HMEX, both of which work in concert to promote glycolysis and inhibit OXPHOS. The result has been consistent across all HMEXs from the six different melanoma cell lines. Our findings reveal a correlation between metabolic function and the presence of metabolically active microRNAs in HMEX that control aerobic glycolysis and OXPHOS of normal fibroblasts. Melanoma exosomes are potentially key players in promoting tumor progression by modulating normal stromal cell metabolism and creating an environment that is conducive to metastasis.

Oxygen Consumption Rate (OCR) pmol/minHADF onlyHADF + HMEX 17.5 μgHADF + HMEX 25 μgHADF + HMEX 50μgHADF + HMEX100μgBasal OCR52.6753.5850.9638.2031.56

Citation Format: Shin La Shu, Cheryl L. Allen, Yunchen Yang, Orla Maguire, Hans Minderman, Arindam Sen, Michael J. Ciesielski, Katherine A. Collins, Peter J. Bush, Martin Morgan, Yun Wu, Richard B. Bankert, Marc S. Ernstoff. Human melanoma exosomes induce metabolic reprogramming in human adult dermal fibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5087.
Abstract 5091: Maintaining long-term stromal depletion therapy for pancreatic cancer
Authors: Grimaldo Sam R. , Khatibeghdami Mahsa
Published: August 2018

A hallmark feature of pancreatic cancer is a dense fibrotic stroma that surrounds the tumor and sequesters blood vessel growth. The lack of blood vessels results in poor perfusion, causing most therapies to fail. Antistromal therapy via targeting the sonic hedgehog (SHH) pathway is a controversial approach to improving drug distribution for pancreatic cancer. While this approach has been shown to be initially effective, over time stroma depletion results in the return of an aggressive cancer. Long-term stromal depletion creates a microenvironment that permits angiogenesis and fuels the return of an aggressive cancer. It is unknown if inhibiting angiogenesis will improve the long-term success of antistromal therapy. We are pursuing two therapeutic strategies that couple an antiangiogenic component with SHH therapy. These therapeutic strategies will be tested in one of the two novel inducible models we have developed, including the standard iKPC model and an SHH-deleted model iKPC-SHH. Preliminary survival analysis shows overall survival of 8 months and 5 months for the iKPC and iKPC-SHH, respectively. Furthermore, increased CD31 blood vessel staining is seen in the iKPC-SHH model, suggesting increased angiogenesis. Using the standard iKPC we will pursue a co-treatment approach with gemcitabine and the SHH inhibitor, itraconazole, to evaluate the effect on median survival. Itraconazole was chosen from a screen of ten SHH inhibitors and found to have a highest efficacy on vascular endothelial cell (IC50 0.02υM). Thereby we expect this molecule to target both the SHH pathway and blood vessels. The second approach we are pursuing is to determine if the antiangiogenic drug, sunitinib, improves median survival in the iKPC-SHH. The iKPC-SHH serves as a surrogate of antistromal therapy and allows for a focused investigation with antiangiogenic compounds. Sunitinib was chosen from the class of antiangiogenic pharmacologics for its direct efficacy on primary pancreatic tumor cells (IC50 2.4υM); thereby it is expected to target blood vessels and tumor cells in vivo. While both treatments approaches are ongoing. current data show that itraconazole co-treatments are living over 7.5 months in the (noninducible) KPC model. Previous reports with an SHH inhibitor in the KPC model showed a survival at approximately 3-4 months. Thereby our initial data show promise that inhibiting angiogenesis can improve the long-term efficacy of antistromal therapy.

Citation Format: Sam R. Grimaldo, Mahsa Khatibeghdami. Maintaining long-term stromal depletion therapy for 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 5091.
Abstract 5101: Characterization of a mouse model using the Sleeping Beauty transposon method to study diffuse intrinsic pontine glioma (DIPG)
Authors: Ravindran Ramya , Mendez Flor M. , Nunez Felipe J. , Koschmann Carl , Dzaman Marta , Pawar Sheeba , Lowenstein Pedro R. , Castro Maria G.
Published: August 2018

Tumors in the brain-stem are difficult to diagnose and treat primarily owing to the location. Diffuse intrinsic pontine glioma (DIPG) is a fatal paediatric brain-stem tumor located in the ventral pons with 100% fatality. These tumors are often treated on the assumption that DIPGs are molecularly similar to adult high grade gliomas; however, they are molecularly distinct. The ACVR1G328V and the H3.1K27M mutations have been seen to recur in DIPGs and we hypothesis that these mutations play an important role in the disease progression. A mutation in the ACVR1 leads to kinase activation and induces downstream signal transduction by phosphorylation of Smads (1/5). H3.1K27M mutations cause genome wide H3K27me3 hypomethylation. We genetically engineered mouse DIPG models using the Sleeping Beauty Transposon (SBT) method to introduce plasmids containing NRASV12/p53-shRNA/ACVR1G328V or NRASV12/p53-shRNA/H3.1K27M into the 4th ventricle of 1-day old pups adjacent to the brain-stem. This generates DIPGs with specific genetic lesions. NRASV12 is a constitutively active oncogene belonging to the RAS family and p53-shRNA is a short hairpin for TP53 tumor suppressor gene. A control tumor model was engineered using plasmids encoding NRASV12 and p53-shRNA. Tumor growth is monitored periodically over time until animals become symptomatic due to tumor burden. At this time point, animals are euthanized and the tumors extracted. We characterized the model for upregulation of phospho-smad 1/5 and markers of neural precursor cells (nestin, Sox2, olig2, GFAP, PDGFRα). In conclusion, we generated a genetically engineered mouse model of DIPG which represents a novel platform to study the molecular pathways underlying disease pathogenesis. An understanding of the functions of mutations that lead up to the disease can help to develop novel therapies for DIPG.

Citation Format: Ramya Ravindran, Flor M. Mendez, Felipe J. Nunez, Carl Koschmann, Marta Dzaman, Sheeba Pawar, Pedro R. Lowenstein, Maria G. Castro. Characterization of a mouse model using the Sleeping Beauty transposon method to study diffuse intrinsic pontine glioma (DIPG) [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 5101.
Abstract 5110: Runx knockout in breast luminal stem/progenitor cells induced precancerous lesion via robust expression of ERα
Authors: Matsuo Junichi , Mon Naing Naing , Yamamura Akihiro , Heng Dede L. , Chuang Linda SH , Osato Motomi , Ito Yoshiaki
Published: August 2018

In breast cancer, 83% of patients were diagnosed as hormone receptor positive, luminal-type, breast cancer (Anderson et al., 2014). Although primary luminal-type breast cancer can be controlled by endocrine therapy, more than 20% of patients have recurrence in 20 years (Pan et al., 2017). Understanding of luminal-type breast cancer would provide benefits to a large number of patients. However, cell of origin and genes responsible for luminal-type breast cancer have not been fully understood. Luminal epithelial cells are considered as origin of breast cancer (Anderson et al., 2014), although not fully proven yet. In addition, extensive genomic analysis of breast cancer patients in TCGA study showed that RUNX genes were downregulated in luminal-type breast cancer: RUNX1 mutation was found in multiple patients, and RUNX3 gene was highly methylated. We previously reported that 270 bp Runx1 enhancer element (eR1) marks hematopoietic stem cell and stomach stem cell (Ng et al., 2010; Matsuo et al., 2017). We also found that this eR1 marks a subset of luminal epithelial cells in mouse mammary gland. Lineage-tracing experiment showed that the eR1+ cells mainly differentiated into luminal cells, but not to basal cells in vivo. These eR1+ cells also contribute to differentiation of alveolar cells in pregnant mice. Interestingly, in vitro lineage tracing by using organoid culture showed that eR1+ cells differentiated into both liminal and basal cells. Runx1 and Runx3 knockout in luminal stem cells showed increase of cell growth, and histology was similar to precancerous lesion such as hyperplasia and ductal carcinoma in situ (DCIS). These lesions showed robust expression of ERα compared with control mice. These results suggested that depletion of Runx in luminal stem cells induces precancerous lesion via robust expression of ERα.

References:

Anderson WF, Rosenberg PS, Prat A, et al. How many etiological subtypes of breast cancer: Two, three, four, or more? J Natl Cancer Inst 2014;106.

Pan H, Gray R, Braybrooke J, et al. 20-year risks of breast-cancer recurrence after stopping endocrine therapy at 5 years. N Engl J Med 2017:1836-46.

Ng CE, Yokomizo T, Yamashita N, et al. A Runx1 intronic enhancer marks hemogenic endothelial cells and hematopoietic stem cells. Stem Cells 2010:1869-81.

Matsuo J, Kimura S, Yamamura A, et al. Identification of stem cells in the epithelium of the stomach corpus and antrum of mice. Gastroenterology 2017:218-31.

Citation Format: Junichi Matsuo, Naing Naing Mon, Akihiro Yamamura, Dede L. Heng, Linda SH Chuang, Motomi Osato, Yoshiaki Ito. Runx knockout in breast luminal stem/progenitor cells induced precancerous lesion via robust expression of ERα [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 5110.
Abstract 5120: Adipocyte-derived lipids mediate melanoma progression via FATP proteins
Authors: Zhang Maomao
Published: August 2018

During melanoma progression, tumor cells come into direct contact with subcutaneous tissues and these advanced lesions portend a poor prognosis. Despite the fact that subcutaneous tissues are largely composed of adipocytes, little is known about the mechanisms by which adipocytes influence melanoma. Using both in vitro and in vivo models, we find that adipocytes increase proliferation and invasion of adjacent melanoma cells. This is accompanied by direct transfer of fatty acids from adipocytes to melanoma cells, which increases tumor cell metabolism and invasion programs. These adipocyte-derived lipids are transferred to melanoma cells through the FATP/SLC27A family of lipid transporters expressed on the tumor cell surface. Among the six related FATP/SLC27A family members, melanomas significantly overexpress FATP1/SLC27A1. Melanocyte-specific FATP1 expression cooperates with BRAFV600E in transgenic zebrafish to accelerate melanoma development, an effect that is similarly seen in mouse xenograft studies. These data demonstrate that adipocytes in the tumor microenvironment can drive melanoma progression through FATP lipid transporters, and represents a new target aimed at interrupting adipocyte-melanoma crosstalk.

Citation Format: Maomao Zhang. Adipocyte-derived lipids mediate melanoma progression via FATP proteins [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 5120.
Abstract 5134: Desacetylvinblastine monohydrazide disrupts tumor vessels by promoting VE-cadherin internalization
Authors: Chen Minfeng , Lei Xueping , Ye Wencai , Zhang Dongmei
Published: August 2018

Vinca alkaloids, the well-known tubulin-binding agents, are widely used for the clinical treatment of malignant tumors. However, little attention has been paid to their vascular disrupting effects, and the underlying mechanisms remain largely unknown. This study aims to investigate the vascular disrupting effect and the underlying mechanisms of vinca alkaloids.

Methods: The capillary disruption assay and aortic ring assay were performed to evaluate the in vitro vascular disrupting effect of desacetylvinblastine monohydrazide (DAVLBH), a derivate of vinblastine, and the in vivo vascular disrupting effect was assessed on HepG2 xenograft model using magnetic resonance imaging, hematoxylin and eosin staining and immunohistochemistry. Tubulin polymerization, endothelial cell monolayer permeability, western blotting and immunofluorescence assays were performed to explore the underlying mechanisms of DAVLBH-mediated tumor vascular disruption.

Results: DAVLBH has potent vascular disrupting activity both in vitro and in vivo. DAVLBH disrupts tumor vessels in a different manner than classical tubulin-targeting VDAs; it inhibits microtubule polymerization, promotes the internalization of vascular endothelial cadherin (VE-cadherin) and inhibits the recycling of internalized VE-cadherin to the cell membrane, thus increasing endothelial cell permeability and ultimately resulting in vascular disruption. DAVLBH-mediated promotion of VE-cadherin internalization and inhibition of internalized VE-cadherin recycling back to the cell membrane are partly dependent on inhibition of microtubule polymerization, and Src activation is involved in DAVLBH-induced VE-cadherin internalization.

Conclusions: This study sheds light on the tumor vascular disrupting effect and

underlying mechanisms of vinca alkaloids and provides new insight into the molecular mechanism of tubulin-targeting VDAs.

Key words: Vascular disrupting agents; vinca alkaloids; desacetylvinblastine monohydrazide; tubulin polymerization; VE-cadherin internalization

Citation Format: Minfeng Chen, Xueping Lei, Wencai Ye, Dongmei Zhang. Desacetylvinblastine monohydrazide disrupts tumor vessels by promoting VE-cadherin internalization [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 5134.