bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1892: Dysfunction of resident memory CD8+ T cells facilitates breast cancer lung metastasis
Authors: Song Erwei , Xing Yue , Su Shicheng
Published: August 2019

Resident memory T cells (TRM) are generally intraepithelially located and manifesting defence potential in diverse existing virus infection models. However, their roles in cancer development, especially metastasis, are poorly understood. Herein, by using a murine model orthotopically inoculated with 4T1 mammary carcinoma cells, we demonstrated the dynamic distribution and functional changes of lung TRM during the formation of lung pre-metastatic niche. Peri-alveolar accumulation of CD8+ T cells, which constitutively express early activation marker CD69 and epithelial-resident marker CD103, is identified. These draining lymph nodes originated and tumor-specific TRM cells, with high expression of IFN-γ and other cytotoxic cytokines, patrol along blood stream and infiltrate in the pre-metastatic lung. IFN-γ coverts alveolar macrophages in the pre-metastatic lung into a immunosuppressive phenotype with high expression of IDO1, which reciprocally compromises the anti-tumor effects of TRM. In conclusion, our study revealed the alteration of lung TRM during the process of metastasis and its interaction with lung pre-metastatic niche, indicating that by finding possible approaches to enhancing TRM function or strengthening local defenses could decrease breast cancer lung-tropic metastasis.

Citation Format: Erwei Song, Yue Xing, Shicheng Su. Dysfunction of resident memory CD8+ T cells facilitates breast cancer lung metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1892.
Abstract 1895: Identification of therapeutic targets in peritumoral microenvironment by single-cell assays using impedance spectroscopy
Authors: Maercker Christian
Published: August 2019

Metastasis and tumor growth are very much dependent on the peritumoral microenvironment. Cells in the peripheral cortex send out growth factors and generate an extracellular matrix which is decisive for infiltration of neoplastic cells and, finally, the formation of a functioning tumor tissue. To figure out specific functions of different cell types and their various interactions in in vitro or in vivo assays, which are possibly relevant for the identification of new drug targets, is a challenge, because we deal with a very complex multifactorial network. Therefore, our goal is to develop test systems with a limited complexity, but which still are informative in terms of molecular tumor development. To reach this, we first have to singularize cells from tumor tissue. Within surgical tissue, tumor core tissue has to be discriminated from surrounding tissue by MRI or immunohistochemistry. Afterwards, microdissected tissue samples have to be dissociated by protease digest. In a next step, the mixture of cells has to be separated into the different cell types by immunopanning and/or FACS sorting. RNAseq helps to identify gene expression profiles specific for individual cells within the tumor compared to the tumor microenvironment. For us the most important parameters are the adhesion and migration activities of the various cell types of different origin on different matrices. To establish a test system as a starting point for a more sophisticated experimental setup to characterize tumorigenic tissues, we firstly isolated mesenchymal stem cells (MSC) from fat tissue or bone marrow, before transfer to microwells covered with electrodes to monitor cell adhesion by electrochemical impedance sensing. We found that pre-coating of the microwells with either collagen type I or IV induced different impedance profiles, corresponding to distinct differentiation pathways which was confirmed by histochemical staining as well as protein and gene expression analyses. In another approach, we used impedance sensing to measure prostate or breast cancer cell migration towards the secretome of mesenchymal stem cells. MSC cell culture supernatant was fractionated by size exclusion and ion exchange chromatography. We identified type I and III collagen, fibronectin and Laminin 421 as extracellular matrix proteins to be the main drivers of rapid cancer cell migration requiring as little as two hours for a full migration response. The challenge now is to adjust this experimental approach to individual cell types isolated from different tumor types and also to different matrices simulating the tumor microenvironment.

Citation Format: Christian Maercker. Identification of therapeutic targets in peritumoral microenvironment by single-cell assays using impedance spectroscopy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1895.
Abstract 1900: A tension-mediated glycocalyx feedback loop promotes glioblastoma
Authors: Kaushik Shelly , Barnes James Matthew , Bainer Russell O. , Sa Jason K. , Woods Elliot C. , Kai Fuiboon , Lakins Jonathon N. , Phillips Joanna J. , Weaver Valerie M.
Published: August 2019

A considerable challenge in glioblastoma (GBM) therapy is treatment resistance and tumor recurrence, resulting in dismal patient prognosis. GBM aggression and recurrence is often associated with a mesenchymal phenotype, increased expression of glycoproteins and the presence of glioma-initiating “stem-like” cells. Using patient-derived xenograft models, and immune competent syngeneic and transgenic glioma mouse models, we show that aggressive mesenchymal-like GBMs, reminiscent of recurrent GBM, are mechanically stiffer, present with a bulky glycocalyx and demonstrate a stem-like phenotype. We report that the more aggressive mesenchymal GBMs show increased mechanical signaling and contractility, and their tumors are surrounded by a stiffer ECM that maintains and even further enhances integrin mechanosignaling. Since a large proportion of these bulky glycoproteins are also stem markers, upregulation of the glycoproteins and their modulators leads to enhanced GBM stem-ness. This was evident both in our mesenchymal models of GBM as well as multiple patient datasets which compared paired primary and recurrent GBM RNA sequencing and protein data. In addition to stem-ness, we show that the mechanically “enhanced” GBMs also foster a differential immune landscape, infiltrated by type 2 macrophages, believed to be pro-tumorigenic and immune evasive. Our findings suggest that there is a dynamic and reciprocal link between integrin mechanosignaling, the GBM immune landscape and a bulky glycocalyx, which promotes a mesenchymal, stem-like phenotype and likely recurrence in GBM patients. Thus, therapeutic strategies to target GBM tissue tension could prevent recurrence, reduce mortality and improve patient outcome.

Citation Format: Shelly Kaushik, James Matthew Barnes, Russell O. Bainer, Jason K. Sa, Elliot C. Woods, Fuiboon Kai, Jonathon N. Lakins, Joanna J. Phillips, Valerie M. Weaver. A tension-mediated glycocalyx feedback loop promotes glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1900.
Abstract 1907: Tenascin-C promotes the activation of mammary fibroblasts to calponin-expressing myofibroblasts
Authors: Katoh Daisuke , Noro Aya , Imanaka-Yoshida Kyoko , Ogawa Tomoko , Yoshida Toshimichi
Published: August 2019

[Introduction] Tenascin-C (TN-C), an extracellular matrix molecules, is highly expressed in cancer stroma. Recent studies have demonstrated that cancer associated fibroblasts (CAFs) expressed TN-C in many tumors, including breast cancer. Here, we clarified the effects of TN-C on phenotype changes of the fibroblasts. [Methods] We examined the cell morphology, expression of CAFs markers, and contractile ability in the fibroblasts after TNC treatment in vitro, using human mammary gland fibroblasts (HMFs) and dermal ones (HDFs). We also investigated the correlation between the expression of the CAF markers and TNC deposition in human breast cancer tissues, using immunohistochemical staining. [Results] TN-C (10μg/mL) induced morphological changes of HMFs from spindle-shaped cells to stellate-shaped cells on plastic as well as on glass coverslips. After the treatment of TN-C, the protein expressions of α-SMA and calponin were significantly increased. Immunofluorescence analysis revealed TN-C increased the actin-stress fiber formation and the expression of α-SMA and calponin. Furthermore, TN-C promoted contraction of collagen gel by HMFs. These results indicated TN-C induces transformation of HMFs into high-contractile myofibroblasts. In human breast cancer tissues, α-SMA and calponin-positive fibroblasts were localized in TN-C-positive cancer stroma. Interestingly, calponin-expressing fibroblasts were not observed in the skin metastatic lesions of breast cancer. In addition, the increased expression of α-SMA and calponin after TN-C treatment in HDFs were significantly lower compared to HMFs in vitro. These results suggested the responses of TN-C on fibroblasts differs according to the residential tissues. Next, we examined the receptor to TNC and signaling pathway for TNC-induced change of HMFs. TN-C increased the colocalization of integrin αv and β1 in focal adhesion. Furthermore, phosphorylation of SMAD2 and SMAD3 were significantly increased and SMAD3 translocated from cytoplasm to nucleus after TN-C addition. Inhibition of TGF-β type I receptor (SB-505124) or SMAD3 (SIS3) suppressed the expression of α-SMA and calponin after TN-C treatment. [Conclusion] TNC could contribute cancer stroma formation characteristics of breast cancer tissues, following induction of phenotypical change to myofibroblasts and enhanced stromal contraction via integrin αvβ1/TGF-β/SMAD signaling. Phenotypic differences between HMFs and HDFs after TN-C administration may be present.

Citation Format: Daisuke Katoh, Aya Noro, Kyoko Imanaka-Yoshida, Tomoko Ogawa, Toshimichi Yoshida. Tenascin-C promotes the activation of mammary fibroblasts to calponin-expressing myofibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1907.
Abstract 1913: Role of glypican 1 in regulating fibroblast activation in the prostate cancer tumor microenvironment
Authors: Kaur Sukhneeraj P. , Lee Hee K. , Arnold Robert D. , Cummings Brian S.
Published: August 2019

Cancer is a heterogeneous collection of neoplastic cells, surrounded by non-malignant, stromal cells, and extracellular matrix (ECM), among other constituents. Cross-talk between tumor and stromal cells forms a dynamic tumor microenvironment (TME) that mediates cell growth, proliferation, and metastasis. The TME can inhibit cancer growth and proliferation during the initial stages of carcinogenesis; however, cancer cells can evade these inhibitory signals, and reshape the TME, making it more amenable to their own growth. This gives rise to cancer-associated stromal cells, including cancer-associated fibroblasts (CAFs). CAFs further modulate the TME by regulating the release of growth signals, cell death, ECM remodeling, angiogenesis, and epithelial-mesenchymal transition. Glypican-1 (GPC1) is a cell surface heparan sulfate proteoglycan (HSPG) that has both pro-mitogenic and pro-metastatic activity. GPC1’s role in regulating fibroblasts and their transformation into CAFs has not been investigated. We studied the ability of prostate cancer cells to alter the TME in vitro by exposing human bone fibroblasts (HS-5 cells) to tumor conditioned media (TCM) isolated from a human neuroendocrine (DU-145) and bone (PC-3) metastatic prostate cancer cell lines and assessed changes in cell morphology and cytotoxicity (using MTT). Exposure of HS-5 cells to TCM from DU-145 and PC-3 cells altered cellular morphology to a more mesenchymal phenotype similar to that seen in active myofibroblasts. TCM also decreased MTT staining in a time-dependent manner. We also determined the effect of GPC1 knockdown (KD) in HS-5 cells on cell morphology, migration and cytotoxicity. Interestingly, we saw phenotypic switching of GPC1 KD HS-5 cells to a myofibroblast like morphology akin to that when treated with TCM. There was no change in cytotoxicity in GPC1 KD HS-5 cells as compared to control cells. However, GPC1 KD enhanced the migration of HS-5 cells in wound healing assay. Migration was also increased in GPC1 KD HS-5 cells upon treatment with TCM. GPC1 KD also increased HS-5 cell contractility significantly. Collectively, these data support the hypothesis that GPC1 plays an inhibitory role in the TME of prostate cancer by regulating the activation of fibroblasts.

Citation Format: Sukhneeraj P. Kaur, Hee K. Lee, Robert D. Arnold, Brian S. Cummings. Role of glypican 1 in regulating fibroblast activation in the prostate cancer tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1913.
Abstract 1918: InnoPanelTM -a well characterized panel of 1000 cancer cell lines for anticancer drug development
Authors: Hao Feng , Liu Changpeng , Peng Hao , Zhou Xiaolin , Fu Yunhui , Hou Qiang , Ning Jinying
Published: August 2019

In vitro cell line assays are broadly used in drug screening to assess drug efficacy, probe drug mechanism, discover and validate predictive biomarkers. High-throughput screen can be performed on cell line collections such as the well-known NCI-60 panel that is consisted of 60 human cancer cell lines developed by the National Cancer Institute (NCI).1Cell lines in such panels are usually well annotated for basic information such as cancer type and subtype, histopathology, and marker gene expression; many experimental parameters are also measured such as doubling time and inoculation density. The utility of cell panels is greatly enhanced with more cell lines that are thoroughly characterized for their genomics by next generation sequencing2-3. To this end, we have developed InnoPanelTM—a panel of 1000 cancer cell lines, including 800 commonly used cancer cell lines, 100 primary cell lines, and 100 engineered cell lines. These cell lines cover all major cancers and majority are genomically characterized with gene expression, mutation and copy number data. Flow cytometry data were also available for engineered cell lines to validate and characterize designed modifications, for example, Siglec-15 overexpression in the CHO-K1 human Siglec-15 cell line. All data are openly available to researchers at our online database InnopediaTM.To make InnoPanelTM-based screening more valuable and accessible to biologists, we developed a bioinformatics pipeline to automate analysis that combines genomics and efficacy data. The utility of such analysis is demonstrated by case studies. InnoPanelTMcan also be customized for a subset of the 1000 cell lines. Subpanel for selected cancer types or cell lines bearing certain features can be constructed. In summary, we developed a well characterized 1000-cell line panel as a valuable resource for researchers working on anticancer drug development.

References1. Shoemaker RH. The NCI60 human tumour cell line anticancer drug screen. Nat Rev Cancer 2006; 6: 813-23.2. Barretina J, Caponigro G, Stransky N, et al. The Cancer Cell Line Encyclopedia enables predictive modelling of anticancer drug sensitivity. Nature, 2012, 483(7391):603.3. Consortium CCLE. Pharmacogenomic agreement between two cancer cell line data sets. Nature, 2015, 528(7580):84-7.

Citation Format: Feng Hao, Changpeng Liu, Hao Peng, Xiaolin Zhou, Yunhui Fu, Qiang Hou, Jinying Ning. InnoPanelTM -a well characterized panel of 1000 cancer cell lines for anticancer drug development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1918.
Abstract 1924: Rapid screening of anti-OX40 antibodies for cancer immunotherapy
Authors: Gan Tian , Li Yanan , Ni Jian , Yang Benny(Yi)
Published: August 2019

OX40, also known as CD134 or TNFRSF4, is a member of the TNFR superfamily. OX40 is a co-stimulatory receptor, and it interacts with its ligand to provide positive signal for T cell activation. OX40 is not expressed on naïve T cells, however its expression goes up after 24 to 72 hours following activation. Disruption of the OX40 pathway led to defective T cell responses, and overexpression of OX40 caused massive immune activation. Due to its importance in keeping immune homeostasis, strategies modulating the OX40 pathway hold great promise in efforts against cancer and autoimmune diseases.

Based on the promising nature of this target, we focused on developing novel OX40 agonists that would effectively stimulate anti-tumor activity. Dozens of high-affinity candidate antibodies were generated using classic hybridoma technology. In order to directly screen for antibodies for anti-tumor effects in animals, we ranked these antibodies in vivo using the drug screening platform based on humanized mouse models. Specifically, we used B-hOX40 mice and implanted syngeneic tumors subcutaneously, followed by treating mice with purified candidate antibodies. We quickly identified several lead antibodies that effectively inhibited tumor growth using this approach, without prior knowledge of their in vitro activities. Their ability to stimulate T cells was confirmed by an engineered reporter line of Jurkat T cells in vitro. Furthermore, we showed that one clone works in concert with Keytruda in a double humanized model of h-OX40 and h-PD-1. We are currently investigating the mechanisms of action underlying these potentially valuable antibodies and carrying out more combination evaluation studies using other available humanized immune-checkpoint mouse models.

Citation Format: Tian Gan, Yanan Li, Jian Ni, Benny(Yi) Yang. Rapid screening of anti-OX40 antibodies for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1924.
Abstract 1949: Mouse model of metastatic colorectal cancer by orthotopic transplantation of patient derived organoids
Authors: Okamoto Takuya , Yaginuma Katsuyuki , Nagayama Satoshi , Yao Ryoji
Published: August 2019

Cancer metastasis proceeds through multiple steps and metastatic colonization is formed by disseminated tumor cells with proliferative capacity in distant tissues. In the analysis of metastasis, macroscopically completed lesions are often used for subjects, which is attributed to the difficulty to capture the early process occurring in living body and the analysis using human specimen is particularly limited. To capture the metastatic process of colorectal cancer (CRC) and to identify cell population involved in it, we produced the mouse model for CRC by orthotopic transplantation of patient derived organoids (PDOs). Unlike ectopic transplantation models, this model precisely recapitulates the metastasis process in human patients. To evaluate the metastasis potentials, the patient matched organoids from the primary, the synchronous liver metastasis and the post-chemotherapy liver recurrence lesion were established. No significant difference in macroscopic morphology or pathogenic mutations were noted among the organoids. The organoids were marked by fluorescent protein and luciferase fusion gene to be visualized the process over time by in vivo imaging system and injected under rectal mucosa on prolapse. Each graft reflected histopathological feature of the surgical specimen. At the results of observation, the lung metastases were frequently occurred in recurrence organoids mouse models, but little occurred in primary or metastasis ones. The lung metastases were considered to be in the early stage of metastasis, because they spread diffusely in each lung lobes, and could not be confirmed macroscopically. They were forming two state; one appeared to be the embolism in blood vessels around the bronchus and others were the micro-metastases in lung tissue. The orthotropic transplantation of patient-matched PDOs provides unique opportunities to explore the metastasis process of CRC.

Citation Format: Takuya Okamoto, Katsuyuki Yaginuma, Satoshi Nagayama, Ryoji Yao. Mouse model of metastatic colorectal cancer by orthotopic transplantation of patient derived organoids [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1949.
Abstract 1962: Visualizing the efficacy of PI3K inhibitor combination treatment on tumor metabolism
Authors: Vaidya Manushka , Sonzogni Olmo , Callahan Cody , Pinto Jocelin , Wulf Gerburg M. , Grant Aaron
Published: August 2019

Purpose: Many types of cancer exhibit an upregulation in glycolysis. Cancer therapies that target tumor metabolism include PI3K inhibitor combinations, which are currently in clinical trials. Real-time metabolic imaging can be possible using hyperpolarization, which is a signal enhancement technique for MRI that increases the signal by a factor of 10,000 or more. The purpose of this work was to determine whether the anti-metabolic effect of such therapies on preclinical breast cancer models could be visualized non-invasively by MRI of hyperpolarized 13C-pyruvate to lactate conversion.

Methods: Experiments were carried out on a 9.4 T animal MRI scanner (Biospec 94/20, Bruker, Billerica MA), and all animal studies were done with IACUC approval. Two models of triple negative BRCA1 mutant breast cancer, either resistant (T1R1, n=4) or sensitive (T2S, n=4) to PARP inhibition, were implanted in the inguinal fat pad of mice. Animals with tumor sizes of approximately 10x10x10 mm3 were then imaged prior to and after four consecutive days of combination treatment with a PI3K inhibitor and PARP inhibitor. In order to visualize the interconversion of pyruvate to lactate in vivo, a bolus of hyperpolarized 13C pyruvate (Hypersense, Oxford Instruments, Oxfordshire UK) with concentration=100 mM and volume=200uL was administered to the mouse via tail vein. 13C MR images were acquired using echo-planar spectroscopic imaging (Flip angle=4 degrees; matrix size=16x16; resolution = 2.5mm; slice thickness=5mm; spectral points=512; spectral width=4kHz; repetitions=32) with a 13C radiofrequency transmit/receive surface coil (Bruker, MA) and overlaid on proton anatomical images obtained using a proton volume coil (78 mm diameter, Bruker, MA). Data were reconstructed (Mathematica, Wolfram) to obtain pixel by pixel time integrated pyruvate and lactate spectra for an axial slice. The area under the curve for both metabolites was calculated for all pixels and summed within the tumor to obtain a lactate to pyruvate ratio for each case. Statistical analysis was carried out using a paired t-test.

Results: For the animals with T2S tumors, the lactate to pyruvate ratio reduced from 1.65 ± 0.44 to 0.84 ± 0.32 (mean ± std. dev.) after treatment with significance p = 0.0378. For animals with T1R1 tumors, the ratio before and after treatment was 0.91 ± 0.44 and 0.93 ± 0.20 (p = 0.9553).

Conclusions: Noninvasive visualization of pyruvate to lactate interconversion can be achieved using 13C MRI, allowing for imaging the efficacy of treatments on cancer metabolism.

Citation Format: Manushka Vaidya, Olmo Sonzogni, Cody Callahan, Jocelin Pinto, Gerburg M. Wulf, Aaron Grant. Visualizing the efficacy of PI3K inhibitor combination treatment on tumor metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1962.
Abstract 197: Molecular interactions of Globo H ceramide with translin-associated factor X (TRAX) underlie its proangiogenic activities
Authors: Yu John , Wang Sheng-Hung , Cheng Jing-Yan , Hung Jung-Tung , Lin Chun-Cheng , Lo Fei-Yun , Lee Chien-Wei , Yu Alice L.
Published: August 2019

Glycosphingolipids (GSLs) are ubiquitous glycolipids in eukaryotic plasma membranes that consist of a polar glycan chain and a hydrophobic sphingosine- containing ceramide tail. Although GSLs display various diversities and functions, the mechanistic details of GSL interactions with membranous or intracellular components are poorly understood. Globo H ceramide (GHCer), a tumor-associated GSL overexpressed in most types of epithelial cancers, is a potential target of cancer immunotherapy. Previously, we showed that GHCer is shed by cancer cells to mediate immune suppression and promote angiogenesis in the tumor microenvironment. At the cellular level, GHCer from cancer cells binds to translin-associated factor (TRAX), with consequent release of PLCB1 from TRAX to trigger Ca2+ mobilization. Herein, we found that Gb5-ceramide (Gb5Cer), the pentasaccharide precursor of GHCer, could not promote angiogenesis nor Ca2+ mobilization. At the molecular level, ELISA and biacore analysis revealed that GHCer binds to TRAX with a distinct dissociation constant of 4.09×10-8 M, while its precursors, galactosyl-, lactosyl-, Gb4-, and Gb5Cer- showed negligible binding to TRAX. Molecular docking and molecular dynamics indicated that the fucose residue of Globo H is an important contributor to a favorable glycan conformation for interaction with TRAX, consistent with the finding that Gb5Cer had no proangiogenic activities. In addition, GHCer competed with recombinant C-terminus region of phospholipase C β1 (PLCB1) for binding to TRAX, thus providing a molecular mechanism for the proangiogenic effect of GHCer on tumor microenvironment. This study provides the first evidence for interaction of TRAX with a non-protein GSL, deciphered distinct regions of TRAX for interacting with glycan and lipid portions of GHCer, and illustrated the crucial contribution of fucose moiety on GHCer for its interaction with TRAX and angiogenic activity.

Citation Format: John Yu, Sheng-Hung Wang, Jing-Yan Cheng, Jung-Tung Hung, Chun-Cheng Lin, Fei-Yun Lo, Chien-Wei Lee, Alice L. Yu. Molecular interactions of Globo H ceramide with translin-associated factor X (TRAX) underlie its proangiogenic activities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 197.
Abstract 1970: Melatonin inhibits osteolytic bone metastasis through interrupting cancer cells-derived osteolytic factor and reduce osteoclast differentiation
Authors: Lai Jyun-Lin , Chang An-Chen , Chen Po-Chun , Tang Chih-Hsin
Published: August 2019

Osteolytic bone metastasis is seen in lung, breast, and prostate cancer and is associated with severe complications such as bone pain, fractures and hypercalcemia. In the bone microenvironment, osteoclasts play an important role in bone remodeling and bone resorption. Metastatic cancer cells may increase osteoclast differentiation and activity by releasing inflammatory cytokines including receptor activator of nuclear factor kappa-B ligand (RANKL), tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6) and IL-8, leading to bone resorption and osteolytic lesions. The endogenous hormone melatonin is known to modulate bone resorption. Increasing evidence suggests that melatonin inhibits cancer cell proliferation, angiogenesis and metastasis. Up until now, the inhibitory role of melatonin in osteolytic bone metastasis has remained unclear. In In vitro study, we observed that melatonin inhibits the differentiation of precursor osteoclasts and promotes apoptosis in mature osteoclasts. We injected mice tibia with luciferase-labeled osteolytic cancer cell lines (lung cancer: A549, prostate cancer: PC-3) to establish bone metastasis. After 4 weeks of intraperitoneal injection with melatonin, IVIS fluorescence signaling indicated that melatonin inhibited tumor growth. X-ray imaging of the bone revealed an improvement in bone resorption after melatonin treatment. In In vitro melatonin also reduced RANKL protein and mRNA expression in the A549 and PC-3 cell lines. Our data indicate that melatonin improves bone mass by inhibiting the expression of cancer-derived osteolytic factors and by promoting apoptosis in mature osteoclasts. Our future investigations will seek to determine the signaling pathway involved in melatonin-regulated RANKL expression in cancer cells. In summary, our findings provide insight into the benefits of melatonin treatment in osteolytic bone metastasis.

Citation Format: Jyun-Lin Lai, An-Chen Chang, Po-Chun Chen, Chih-Hsin Tang. Melatonin inhibits osteolytic bone metastasis through interrupting cancer cells-derived osteolytic factor and reduce osteoclast differentiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1970.
Abstract 1974: Autocrine TIMP-1-mediated chemoresistance via induction of IL-6 in NSCLC
Authors: Xiao Wei , Rojiani Amyn M. , Gupta Sampa Ghoshal , Rojiani Mumtaz V.
Published: August 2019

Our previous studies have confirmed the well-defined antiapoptotic functions of TIMP-1. We have shown that overexpressing TIMP-1 in NSCLC cells increases cell survival with activation of the ERK pathway via phosphorylation of BAD at serine 112. Knocking down TIMP-1 in NSCLC cells increased the levels of miR-125a-5p with concomitant upregulation in apoptosis. These studies beg the question- does modulation of TIMP-1 affect chemotherapy in NSCLC? To address this question we examined two cell lines, A549 and H460, and their specific shRNA-mediated TIMP-1 gene knockdown clones. We found that TIMP-1 mediated chemoresistance against two front line chemotherapeutic agents (Gemcitabine and Cisplatin) as determined by increased apoptosis in the KD clones. In order to determine any contribution of cytokine activity towards this antiapoptotic function of TIMP-1, we carried out a cytometric bead array assay. The results of this screening assay which included potential inflammatory cytokines and growth factors, indicated that IL-6 plays a role in mediating the chemoresistance in TIMP-1 expressing NSCLCs. We then determined endogenous IL-6 levels in our array of NSCLC cell lines and found a direct correlation of IL-6 levels with TIMP-1 levels. It is important to note that IL-6 expression is precisely regulated with TIMP-1 production at both protein and mRNA levels. The effect of TIMP-1 on IL-6 levels is more pronounced under hypoxic versus normoxic conditions. Importantly, IL-6 expression is upregulated by addition of exogenous human recombinant TIMP-1 in a time- and dose- dependent manner. Blocking of TIMP-1 signaling with specific neutralizing antibodies delayed the IL-6 upregulation. Moreover, addition of rhTIMP-1 or rhIL-6 rescued apoptosis induced by chemotherapeutic agents albeit to a limited extent. Although early in the investigation, our studies suggest that interventions impacting the TIMP-1/IL-6 axis in NSCLCs may provide an efficient target to boost effects of clinical chemotherapy.

Citation Format: Wei Xiao, Amyn M. Rojiani, Sampa Ghoshal Gupta, Mumtaz V. Rojiani. Autocrine TIMP-1-mediated chemoresistance via induction of IL-6 in NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1974.
Abstract 1975: Multivariate exome analysis identifies CXCR5-CXCL13 signaling as a key driver in early breast cancer development and prognosis
Authors: Dill Courtney D. , Griffen Tiara , Young Corey , Carey Kaylin , Lillard James W.
Published: August 2019

Breast cancer (BrCa) is the most frequently diagnosed cancer and the second leading cause of cancer-related death among women worldwide despite new developments in prevention and treatment. More recently, rapid increases in BrCa incidence have been reported in countries of the Asia-Pacific region. Age of diagnosis in Asian BrCa patients is much younger than age of diagnosis in western countries, such as the United States, and Asian early BrCa patients tend to be pre-menopausal, whereas only 15-30% of western BrCas are pre-menopausal. The mechanisms responsible for early BrCa remain unknown. Furthermore, it remains controversial whether early breast cancer has unique tumor biology. The goal of this study was to characterize the molecular phenotype of early BrCa in the context of CXCL13, CXCR5 and associated gene expression. We hypothesized that CXCR5-CXCL13 signaling contributes to the aggressive phenotype of early BrCa. A bioinformatic approach was used to aid in characterizing this condition. Our patient cohort data (50 females of Korean descent, age ≤ 35 years) was obtained from The International Cancer Genome Consortium (ICGC). DESeq analysis was used to identify genes differentially expressed among normal tissue and primary tumor groups. Weighted Gene Network Co-expression (WGCNA) analysis was carried-out to identify gene networks associated with factors influencing BrCa prognosis. Finally, canonical pathway, upstream regulator, and gene interaction analysis was conducted using Ingenuity Pathway Analysis. Our data show CXCL13, CXCR5, and associated genes (LTA, LTB, IL10, IL27RA, TNFRSF13B, TNFRSF17, and TNFRSF9), which have been previously shown to drive tertiary lymphoid structure (TLS) formation, were significantly associated with the molecular phenotype of early BrCa in our patient cohort. Taken together, our findings suggest TLSs contribute to the tumor immune microenvironment and contribute to early BrCa progression.

Citation Format: Courtney D. Dill, Tiara Griffen, Corey Young, Kaylin Carey, James W. Lillard. Multivariate exome analysis identifies CXCR5-CXCL13 signaling as a key driver in early breast cancer development and prognosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1975.
Abstract 1979: Interaction of atorvastatin and CX3CR1-fractalkine in androgen-dependent prostate cancer cells: Effect on PI3K pathway
Authors: Al-Husein Belal
Published: August 2019

Prostate cancer (PC) management lacks any targeted therapies. Statins have shown anticancer effects against PC in both in vitro and in vivo studies. CX3CR1 is a receptor expressed by many cancer cells, including PC. In this research, we studies the interaction between atorvastatin and CX3CR1/fractalkine on androgen-dependent PC cell line 22Rv1 vs. normal prostatic cells, RWPE1. Experimental settings were based on testing effect of atorvastatin in presence of fractalkine or by removing its effect using fractalkine neutralizing antibodies. MTT proliferation assay and adhesion assay, together with testing the expression of CX3CR1, p-AKT and p-GSK-3 were used. Results showed that fractalkine partially blunts the effect atorvastatin on proliferation and adhesion of PC cells. Similar effects were not noticed on normal prostatic cells. Western blotting showed a dose-dependent reduction in the expression of CX3CR1, p-akt and p-GSK-3 in presence of fractalkine, that was further enhanced when fractalkine was neutralized using specific antibodies. Results suggest the deprivation of fractalkine as a synergistic mechanism to enhance effects of atorvastatin and probably other anticancer agents.

Citation Format: Belal Al-Husein. Interaction of atorvastatin and CX3CR1-fractalkine in androgen-dependent prostate cancer cells: Effect on PI3K pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1979.
Abstract 1992: Mesenchymal stem cell-derived interleukin-28 can drive the selection of apoptosis resistant bone metastatic prostate cancer
Authors: Mcguire Jeremy J. , Cook Leah , Frieling Jeremy , Muhammad Ayaz , Lawrence Harshani , Lawrence Nicholas , Lynch Conor
Published: August 2019

Bone metastatic prostate cancer is painful and incurable. We believe that understanding the interactions of tumor cells and the surrounding stromal cells may provide new therapeutic options. Bone metastatic prostate cancer promotes mesenchymal stem cell (MSC) recruitment and differentiation into osteoblasts, leading to the clinically often-observed osteogenic phenotype. However, the reciprocal roles of bone- marrow derived MSCs on prostate cancer cells are less explored. Here, we discovered that MSCs significantly suppress prostate cancer growth in vitro and in vivo. Mechanistically, we show that MSC-derived interleukin-28 (IL-28) promotes prostate cancer cell apoptosis via the IL-28 receptor alpha (IL-28Rα). However, chronic exposure to MSCs yields prostate cancer cell populations that are resistant to IL-28 induced apoptosis and therapeutics such as docetaxel. In vivo, we observed that MSC selected prostate cancer cells rather than being suppressed, grow at a significantly enhanced rate in bone. The increase in apoptosis resistance is accompanied by a shift in downstream signaling of the IL-28Rα from STAT1 to STAT3. Reduction of STAT3 levels or pharmacological inhibition (STI-201) significantly reduces the growth of MSC selected prostate cancer cells in vitro and in vivo. Collectively, these data shed light on how MSCs in the bone marrow microenvironment initially protect against prostate cancer cell establishment but in doing so, help select for aggressive, apoptosis resistant bone metastatic prostate cancer cells that are more aggressive. Given that the majority of prostate to bone metastases are positive for STAT3 expression and activity, our data provide rationale for the therapeutic targeting of STAT3 in bone metastatic prostate cancer.

Citation Format: Jeremy J. Mcguire, Leah Cook, Jeremy Frieling, Ayaz Muhammad, Harshani Lawrence, Nicholas Lawrence, Conor Lynch. Mesenchymal stem cell-derived interleukin-28 can drive the selection of apoptosis resistant bone metastatic prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1992.
Abstract 1999: As4O6 suppresses triple-negative breast cancer via inhibiting EGFR/mTOR mediated EMT in vitro and in vivo
Authors: Bae Illju , Lian Zenglin , Liu Ziyao , Wu Zhaoyan
Published: August 2019

Objective:The present study aimed to verify the anti-metastasis effects and its mechanisms of As4O6 on triple-negative breast cancer(TNBC)in vitro and in vivo.

Methods:MTT assay was used to evaluate cell proliferation. Wound healing assay and the invasion assay were carried out to investigate cell migration and invasion. The xenograft zebrafish model and orthotopic xenograft nude mouse model derived from MDA-MB-231-Green fluorescent protein (GFP) cells was established to analysis the anti-metastasis effect of As4O6in vivo. Western blot and immunohistochemistry were performed to analysis the metastasis-related proteins expression.

Result: MTT assay showed cell proliferation of TNBC cell MDA-MB-231 was inhibited in a dose-dependent manner by As4O6 treatment. Also, As4O6 suppressed cell migration and invasion by a Wound healing assay and the invasion assay, resulted in the reduction of MMP-2 and MMP-9 expression, especially for the inhibition of EGFR and mTOR phosphorylation. Meanwhile, As4O6 suppressed cancer metastasis in xenograft zebrafish model. The number of metastatic lesion in mouse lung was significantly reduced by the treatment of As4O6 in the orthotopic xenograft nude mouse model. The expression of p-mTOR, p-EGFR and EMT-related proteins such as Vimentin and E-cadherin were significantly reversed in As4O6 treated mouse lung tissues than that of non-treatment group. Furthermore, As4O6 treatment suppressed the expression of angiogenesis related proteins VEGF-a, VEGFR, CD34 and MMP2/9 in mouse lung tissues.

Conclusion:Thus, the data suggests that As4O6suppresses triple-negative breast cancer metastasis via inhibiting EGFR/mTOR mediated EMT in vitro and in vivo. As4O6 might be served as a potential useful agent for TNBC metastasis in the future.

Citation Format: Illju Bae, Zenglin Lian, Ziyao Liu, Zhaoyan Wu. As4O6 suppresses triple-negative breast cancer via inhibiting EGFR/mTOR mediated EMT in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1999.
Abstract 20: Pharmacology and metabolism of GLL398, an oral selective estrogen receptor degrader for endocrine therapy of breast cancer
Authors: Guo Shanchun , Zhang Changde , Mottamal Madhu , Hossain Ahamed , Liu Jiawang , Liu Jiawang , Wang Guangdi
Published: August 2019

Selective estrogen receptor degrader (SERD) has proven clinically effective in treating advanced or metastatic breast cancer since the approval of fulvestrant by FDA in 2002. Recent expansion of indications as a first line monotherapy and as combination therapy with CDK4/6 inhibitors further demonstrates its clinical utility as an efficacious breast cancer endocrine regimen. However, the poor pharmacokinetic properties of fulvestrant and its injection-only administration route has driven continued efforts to develop orally bioavailability SERD that could potentially improve clinical response to SERD treatment. GLL398, a boron-modified GW5638 analog, showed superior oral bioavailability while retaining both antiestrogenic activity and ER degrading efficacy at a potency level comparable to the more active metabolite of GW5638, GW7604. Here we report further studies on the pharmacology, pharmacokinetics, and metabolism of GLL398. Consistent with GLL398’s robust activities in breast cancer cells that are tamoxifen resistant or express constitutively active, mutant ESR1 (Y537S), it was found to bind the mutant ERY537S at a high affinity. Molecular modeling of the binding mode of GLL398 to ER also found its molecular interactions consistent with the experimentally determined high binding affinity towards WT ER and ERY537S. To test if the superior oral bioavailability can be translated to potent efficacy in vivo, mice bearing MCF-7 derived xenograft breast tumors and patient derived xenograft (PDX) tumors harboring ERY537S were treated with GLL398 which potently inhibited tumor growth in mice. Finally the metabolism of GLL398 was also investigated to elucidate the biotransformation of the drug and its potential contribution to the superior oral bioavailability over GW7604.

Citation Format: Shanchun Guo, Changde Zhang, Madhu Mottamal, Ahamed Hossain, Jiawang Liu, Jiawang Liu, Guangdi Wang. Pharmacology and metabolism of GLL398, an oral selective estrogen receptor degrader for endocrine therapy of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 20.
Abstract 2002: Computational drug repositioning identifies bortezomib as a novel metastatic inhibitor of lung cancer
Authors: Kwon Ok-Seon , Lee Haeseung , Kim Wankyu , Cha Hyuk-Jin
Published: August 2019

Although many molecular targets for cancer therapy have been discovered, they often show poor druggability, which is a major obstacle to develop targeted drugs. As an alternative route to drug discovery, we adopted an in silicodrug repositioning (in silicoDR) approach based on large-scale gene expression signatures, with the goal of identifying inhibitors of lung cancer metastasis. Our analysis of clinicogenomic data identified GALNT14, an enzyme involved in O-linked N-acetyl galactosamine glycosylation, as a putative driver of lung cancer metastasis leading to poor survival. To overcome the poor druggability of GALNT14, we leveraged Connectivity Map approach, an in silicoscreening for drugs that are likely to revert the metastatic expression patterns. It leads to identification of bortezomib (BTZ) as a potent metastatic inhibitor, bypassing direct inhibition of poorly druggable target, GALNT14. The anti-metastatic effect of BTZ was verified in vitroand in vivo. Notably, both BTZ treatment and GALNT14knockdown attenuated TGFβ-mediated gene expression and suppressed TGFβ-dependent metastatic genes, suggesting that BTZ acts by modulating TGFβsignaling. Taken together, these results demonstrate that our in silicoDR approach is a viable strategy to identify a candidate drug for undruggable targets, and to uncover its underlying mechanisms.

Citation Format: Ok-Seon Kwon, Haeseung Lee, Wankyu Kim, Hyuk-Jin Cha. Computational drug repositioning identifies bortezomib as a novel metastatic inhibitor of lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2002.
Abstract 2007: Validation of a preclinical model for bone metastatic triple negative breast cancer
Authors: Zdrojewska Justyna , Suominen Mari I. , Fagerlund Katja M. , Halleen Jussi M. , Bernoulli Jenni , Zimmermann Johann
Published: August 2019

Metastatic breast cancer (MBC) is the most advanced stage of breast cancer, diagnosed as stage IV. Typically, MBC metastases occur in the bones, liver, brain, and lungs. Bone metastases cause significant morbidity and mortality in late-stage breast cancer patients. At present, there is no efficient curative or preventive treatment for bone metastases. Therefore, establishment and validation of well characterized in vivo models of breast cancer bone metastasis is of utmost importance. The aim of this study was to validate an intracardiac model of bone MBC utilizing GFP tagged triple negative (estrogen receptor; ER, progesterone receptor; PR and human epidermal growth factor receptor 2; HER2 negative) human breast cancer cells, which could be used to study efficacy of new potential treatments. Female athymic nude mice (Hsd: Athymic Nude-Foxn1nu) were used in this study. Bone seeking MDA-MB-231(SA)-GFP human breast cancer cells were inoculated into the left cardiac ventricle of 4-6 weeks old female mice. Allocation to treatment groups (n=15) was performed by stratification procedure based on animal weight prior to the beginning of the study. To validate the model, the current standard-of-care (SOC) treatment (Paclitaxel) was administered between study days 13-17 at the therapeutic dose. Serum bone biomarker TRACP5b was measured before cancer cell inoculation and at sacrifice. Planar X-ray of hind limbs and ex vivo fluorescence imaging of the whole body were performed at sacrifice. The observed tumor take rate was 100%. 33% of mice in the vehicle group were cachectic, but mice treated with SOC exhibited only mild and temporary weight loss. Tumors were observed mainly in skeletal sites but also in soft tissues. The SOC treatment decreased both total and skeletal tumor burden. Based on X-ray images, tumor induced osteolytic bone lesions were smaller in the SOC treated group compared to vehicle though no differences were observed in total bone lesion area. Serum TRACP5b was increased in the vehicle group as a result of osteolytic lesions, and decreased with the SOC treatment. In conclusion, an intracardiac model of MBC was successfully validated with the SOC treatment paclitaxel that efficiently inhibited disease progression. Therefore, this intracardiac model provides a promising tool for testing new treatments against bone and soft tissue MBC in vivo.

Citation Format: Justyna Zdrojewska, Mari I. Suominen, Katja M. Fagerlund, Jussi M. Halleen, Jenni Bernoulli, Johann Zimmermann. Validation of a preclinical model for bone metastatic triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2007.