bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4509: Overexpression of CADM1 enhances malignant features of small-cell lung cancer
Authors: Funaki Toko , Ito Takeshi , Murakami Yoshinori
Published: August 2019

Small-cell lung cancer (SCLC) is one of the representative intractable cancers with the 5-year survival of less than 10 %. Novel approaches to the treatment of SCLC are required because recurrence arises inevitably after the initial chemotherapy. CADM1 was originally identified as a tumor suppressor of non-small cell lung cancer (NSCLC) which encodes a member of immunoglobulin superfamily cell adhesion molecules. CADM1 consists of an extracellular domain with three immunoglobulin-like loops, a trans-membrane domain and a short cytoplasmic domain containing a protein 4.1-binding motif (4.1BM) and a PDZ-II-binding motif. We have reported that CADM1 is highly expressed in 90% of human SCLC cell lines and promotes subcutaneous tumor formation of SCLC cells in nude mice. In this study, we performed clinicopathological evaluation of CADM1 in SCLC and analyzed the molecular mechanisms of malignant progression of SCLC by overexpressing CADM1. Firstly, we performed immunohistochemical analysis of CADM1 expression in high-grade neuroendocrine tumor (HGNEC) of the lung, including SCLC and large cell neuroendocrine carcinoma (LCNEC). We found that CADM1 protein was expressed in 75% (33/44) of SCLC tumors and correlated with the nodal involvement in lung HGENC. To understand pathological significance of CADM1 overexpression in SCLC, we introduced a full-length CADM1 and its mutants in the extracellular or intracellular domain into SCLC cells, SBC5, lacking endogenous CADM1 expression and assessed their malignant features in comparison with control SBC5 cells. CADM1 expression significantly enhanced colony formation in soft agar and subcutaneous tumor formation in nude mice of SBC5, whereas a mutant in 4.1BM did not. Then, we screened chemical library of 170 small molecules of known function to identify compounds which inhibit anchorage-independent growth of SCLC cells enhanced by CADM1. Inhibitors of PI3K, Akt and PKA significantly suppressed cell growth of SBC5, suggesting that CADM1 activates PI3K/Akt and PKA signaling pathways through its 4.1BM and promotes anchorage-independent growth of SCLC cells. We also found that orthotopic injection of NCI-N417 cells overexpressing CADM1 into the lung of nude mice significantly enhanced their metastasis to mediastinal lymph nodes. These results suggest that CADM1 enhances tumor growth and metastasis of SCLC cells probably by activating PI3K/Akt and PKA pathways through its binding proteins at 4.1BM.

Citation Format: Toko Funaki, Takeshi Ito, Yoshinori Murakami. Overexpression of CADM1 enhances malignant features of small-cell 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 4509.
Abstract 4511: Prostate cancer-specific enrichment of TP53 missense mutations elicits differential, context-dependent biochemical and biologic outcomes
Authors: McCann Jennifer J. , Vasilevskaya Irina , PoudelNeupane Neermala , Dean Jeffry , Mandigo Amy , Leeuw Renee De , McNair Chris , Schiewer Matthew , Knudsen Karen
Published: August 2019

Prostatic adenocarcinoma (PCa) remains the second leading cause of cancer death in men in the United States. While organ-confined disease can be successfully treated with surgery or radiation, patients with advanced disease undergo androgen deprivation therapy (ADT), which targets the androgen receptor (AR) signaling axis. This approach effectively treats the disease as PCa cells are exquisitely dependent on AR signaling for proliferation and survival. Unfortunately, cancer progression resumes after 2-3 years and results in a lethal stage of disease, termed castration-resistant prostate cancer (CRPC). While PCa typically demonstrates low overall mutational burden, TP53 is frequently mutated in both primary and advanced disease. These mutations are most commonly missense mutations in the DNA binding domain of TP53, and can occur in the presence or absence of a wildtype TP53 allele. Interestingly, specific enrichment of certain TP53 mutations frequently occurs in PCa when compared to other cancers, and similar to many other cancers, these mutations occur in the presence or absence of wildtype TP53. Using hormone therapy sensitive and CRPC cells, a panel of cell lines was generated to model these mutations in the presence or absence of wildtype TP53 expression. Defining the TP53 missense mutant-sensitive transcriptomes demonstrated context dependent, differential gene expression between TP53 missense mutants, related to the expression of wildtype TP53 in those cells. This also applied to cistrome analysis, illustrating an expansion of the p53 cistrome upon expression of mutant TP53. Furthermore, expression of p53 mutants also elicited context dependent effects on canonical p53 functions, thereby modulating distinct, downstream biological outcomes. Consequently, these data define the distinct and context dependent roles that mutant TP53 plays in PCa progression.

Citation Format: Jennifer J. McCann, Irina Vasilevskaya, Neermala PoudelNeupane, Jeffry Dean, Amy Mandigo, Renee De Leeuw, Chris McNair, Matthew Schiewer, Karen Knudsen. Prostate cancer-specific enrichment of TP53 missense mutations elicits differential, context-dependent biochemical and biologic outcomes [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 4511.
Abstract 4518: IL-4/IL-13 stimulated tumor-associated macrophages enhance breast cancer cell invasion through Rho-GTPase signaling
Authors: Little Andrew C. , Pathanjeli Pragathi , Wu Zhifen , Bao Liwei , Goo Laura , Yates Joel A. , Merajver Sofia D.
Published: August 2019

The high metastatic potential of inflammatory and other aggressive breast cancer is the major determinant of mortality. Recently, it has become clear that signals from the tumor microenvironment (TME) harbor the capacity to enhance cancer cell dissemination. Of the various cells that comprise the TME, macrophages are the most abundant in solid tumors. Therefore, we aimed to better understand the relationship between macrophage-breast cancer cell crosstalk. Intriguingly, in our previous work we showed that inflammatory breast cancer (IBC) cells are hyper-migratory in response to macrophage conditioned media. Additionally, we found this process is regulated by the Rho-GTPase, RhoC. Recent advances in tumor associated macrophage (TAM) biology have led to the understanding that TAM populations in the TME are diverse, primarily due to directional differentiation promoted by unique secreted components from the various cells in the TME. Therefore, this study aimed to understand which subpopulation of M2-like TAMs had the greatest impact on breast cancer cell aggressiveness. Our data show that stimulation of the triple negative breast cancer (TNBC) cell model, MDA-MB-231, and the IBC TNBC cell line, SUM-149, with media extracts from IL-4/IL-13 stimulated M2 macrophages (M2a) elicit the strongest migratory and invasive responses. Subsequently, we stimulated RhoA or RhoC knockout (CRISPR) MDA-231 and SUM-149 cells with M2a conditioned media to address their role in regulating migratory/invasive responses. In brief, we find that RhoA and RhoC, in part, regulate M2a-TAM induced responses. Secretome analysis of M2a-TAM conditioned media reveals high levels of vascular endothelial growth factor (VEGF) and chemokine (C-C motif) ligand 18 (CCL-18). Functional studies suggest that VEGF and CCL-18 synergistically enhance cellular invasiveness. Moreover, pretreatment with the ROCK inhibitor Y-27632 or GSK429286A drastically inhibited VEGF, CCL-18, or M2a induced migratory responses. These data suggest that the Rho-GTPases regulate M2a-mediated breast cancer cell invasion and potentially offer a novel approach for the treatment of metastatic breast cancer through potential prevention of progression.

Citation Format: Andrew C. Little, Pragathi Pathanjeli, Zhifen Wu, Liwei Bao, Laura Goo, Joel A. Yates, Sofia D. Merajver. IL-4/IL-13 stimulated tumor-associated macrophages enhance breast cancer cell invasion through Rho-GTPase signaling [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 4518.
Abstract 4520: Lymphatic vessels regulate exosome trafficking from tumors
Authors: Maillat Lea , Potin Lambert , Kilarski Witold W. , Swartz Melody A.
Published: August 2019

Prior to metastasis, tumor-draining lymph nodes and distant organs are altered by a number of tumor-derived factors that help to both suppress host immunity as well as form an environment that supports metastatic tumor cells, i.e., a pre-metastatic niche. Recent studies suggest that pre-metastatic niches are induced by vesicles, including exosomes, secreted by tumor cells and cells of the tumor microenvironment. Here, we investigated the contribution of lymphatic vessels, particularly the lymphatic endothelial cells (LECs), in the active regulation of tumor exosome production and trafficking. To track exosomes, we purified and fluorescently labeled exosomes from in vitro cultures of the mouse melanoma cell line B16 F10 and of the human melanoma cell line Me260LN. Upon injection into the mouse ear dermis, fluorescent exosomes were observed within minutes in draining lymphatics, but not in surrounding blood vessels. Using the B16 F10 melanoma model overexpressing the lymphangiogenic growth factor VEGF-C, we found that upon intratumoral injection of exosomes, LECs within the tumor microenvironment abundantly take up exosomes, and at higher levels than blood endothelial cells, delivering exosomes to draining lymph nodes and eventually to the blood. In transgenic mice lacking dermal lymphatics, exosomes injected intratumorally were absent in the lymph nodes and present in far lower amounts in the blood compared to those injected into wild type mice. Additionally, while the density of endogenous blood exosomes increased upon tumor growth in wild type mice, no significant change in blood exosome density was observed in transgenic mice. Using an in vitro model where LECs are seeded onto porous inserts, we found that LEC transport of tumor exosomes occurs preferentially from their basal to apical side, and that some of these exosomes were taken up and stored intracellularly. Together, these findings suggest that lymphatic vessels constitute a major route of tumor exosome distribution to the systemic circulation and that LECs actively regulate exosome transport by active uptake.

Citation Format: Lea Maillat, Lambert Potin, Witold W. Kilarski, Melody A. Swartz. Lymphatic vessels regulate exosome trafficking from tumors [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 4520.
Abstract 4541: Malignant and non-malignant cells from primary tumor and lymph node metastasis ecosystems show different behavior for patient-associated protein signatures in head and neck cancer
Authors: Busso-Lopes Ariane F. , Rivera César , Mello Barbara P. , Villa Luisa L. , González-Arriagada Wilfredo , Leme Adriana F. Paes
Published: August 2019

Although lymph node metastasis is the main prognostic factor in patients with head and neck squamous cell carcinoma (HNSCC), the molecular signals involved in this process are poorly characterized and no molecular markers are currently used in clinical practice. Recently, cancer is being viewed as an ecosystem, in which tumor cells cooperate with each other and host cells in their microenvironment. Herein, we mapped the proteome of primary tumor and matched lymph node ecosystems, isolating malignant and non-malignant cells, to better understand the mechanisms underlying lymph node metastasis. For that, we used formalin-fixed paraffin-embedded (FFPE) primary tumors and paired lymph nodes from 27 HNSCC patients positive (N+, n=13) and negative (N0, n=14) for lymph node metastasis using laser microdissection combined with mass spectrometry-based proteomics. An average of 2,266 ± 216 proteins was identified in primary tumor and lymph node ecosystems. A hundred and five differentially abundant proteins from malignant cells in the primary site (N+ vs N0) were mainly involved in RNA stability and metabolism, while inflammation and splicing processes, respectively, were overrepresented in N+ vs N0 non-malignant cells signatures from primary tumor (N=33) and lymph node (N=13) sites (Student's t-test; P-value≤0.05), indicating particular lymph node metastasis signatures for each cell population. Interestingly, the protein pattern of malignant cells from the primary site and matched lymph nodes showed an individual patient-specific metastasis profile through hierarchical clustering analysis; however, the profile of non-malignant cells does not represent patient-specific subpopulations and grouped according to the site of isolation (primary tumor and lymph node microenvironment). Our data reveal potential protein signatures associated with prognosis in HNSCC, as well as a non-specificity of protein patterns from host non-malignant cells that can be related with the immunosurveillance failure. Financial support: FAPESP (Process number 2015/19191-6).

Note: This abstract was not presented at the meeting.

Citation Format: Ariane F. Busso-Lopes, César Rivera, Barbara P. Mello, Luisa L. Villa, Wilfredo González-Arriagada, Adriana F. Paes Leme. Malignant and non-malignant cells from primary tumor and lymph node metastasis ecosystems show different behavior for patient-associated protein signatures in head and neck 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 4541.
Abstract 4565: The functional role of atypical chemokine receptor 1 in immune cell regulation of breast cancer
Authors: Jenkins Brittany D. , Martini Rachel , Gardner Kevin , Monteil Michele , Deeb Dorrah , Newman Lisa , Davis Melissa
Published: August 2019

Breast cancer (BC) is a heterogeneous disease that leads to varied molecular subtypes and distinct clinical outcomes. Tumor heterogeneity, which is in part influenced by genetic ancestry, contributes to racial disparities in BC. This disparity has persisted for over 30 years, despite improvements in detection, screening, and treatment methods. When investigating race-specific differences in the breast tumor microenvironment (TME), we believe that chemokine receptors, such as atypical chemokine receptor 1 (ACKR1/DARC), play an integral role in regulating chemokine and immune cell migration in circulation and the TME, ultimately influencing tumor progression. ACKR1/DARC specifically plays a role in ancestry-related differences in BC due to an African-specific ancestral allele (“Duffy-null” mutation) that is fixed in Sub-Saharan African populations, and present in 60-70% of African-Americans. Those that harbor the mutation do not express ACKR1/DARC on erythrocytes, ultimately affecting immune cell migration in these populations only. To better understand the effects of this variant in circulation and the TME, we performed immunohistochemistry (IHC) on breast tumors from a cohort of patients with matched blood samples. From our IHC analysis, we quantified levels of ACKR1/DARC, in addition to levels of target pro-inflammatory chemokines, and distinct immune populations of T-cells and macrophages. In addition, individuals were genotyped to determine Duffy-null status, and a correlative Luminex multiplex assay was performed on patient plasma to determine concentrations of pro-inflammatory chemokines in circulation. We determined that those with low expression of ACKR1/DARC in tumors exhibited a unique signature of immune cell infiltrates that would encourage a more aggressive TME. In addition, we observed variable levels of circulating chemokines, where those with the Duffy-null mutation exhibited significantly decreased levels of CCL2. Overall, our analyses support the hypothesis that tumor-specific DARC/ACKR1 plays a vital role in immune cell regulation in women with BC.

Citation Format: Brittany D. Jenkins, Rachel Martini, Kevin Gardner, Michele Monteil, Dorrah Deeb, Lisa Newman, Melissa Davis. The functional role of atypical chemokine receptor 1 in immune cell regulation 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 4565.
Abstract 4596: Synthesis and biological evaluation of small molecules inhibitors targeting heterotrimeric Gαi2 protein
Authors: Caggia Silvia , Garrett Autumn S. , Kumar Aditi , Tapadar Subhasish , Oyelere Adegboyega K. , Venugopal Smrruthi Vaidegi , Khan Shafiq A.
Published: August 2019

Heterotrimeric G-proteins are ubiquitously expressed in cells, and they transduce signals from activated G-protein coupled receptors. It is well known that these proteins are differentially expressed in cancer, thereby activating signals that lead to different biological functions such as proliferation and cell motility. Previously, we have shown that Gαi2 protein is essential for cell migration and invasion in prostate cancer cell lines, and its action is downstream of PI3-kinase/Rac1 activation. We have also shown that in PC3 prostate cancer cells, pre-treated with Pertussis toxin (PTX), a well know inhibitor of Gαi/0, resulted in the attenuation of TGFβ1- and oxytocin-induced migration and PI3-kinase activation, without affecting EGF-induced PI3-kinase activation and cell migration. Here, we synthesized new small molecules inhibitors, GDIs, capable of preventing nucleotide exchange and subunit activation. Compounds #29 and #46 are analogs of the lead compound, lacking the thiophene OH-group and with a thiol- to N-methyl amino-group substitution, respectively. Compound #35, a methyl ether derivative of #46, is designed to test the effect of the modification to the phenolic group on Gαi2 inhibition activity. We pretreated PC3 cells with the small molecules and then stimulated with EGF and oxytocin to activate Gαi2 subunit. Immunoprecipitation of the active Gαi and western blot for Gαi2 showed a decrease in the amount of active Gαi2 that was pulled down compared with the controls. Using cell migration assay, we observed that #29 and #46, both at 10 μM, caused inhibition of migration in different prostate cancer cell lines in response to EGF treatments. In contrast, #35 is inactive in this assay at the same concentration. To determine if the effects of Gαi2 are the same in other cancer cells, we first knockdown Gαi2 protein expression in breast and ovarian cancer cells and performed migration assays. The knock-down of endogenous Gαi2 attenuated cell migration, compared to the control cells. We also performed migration assays in the same cells using compound #29 at 10 μM and observed significant inhibition of migratory behavior in response to EGF- and FBS stimuli compared to the controls. We conclude that the small molecules we used in this study may be considered as potential therapeutic tools, inhibiting the migratory capability of metastatic cells.

Citation Format: Silvia Caggia, Autumn S. Garrett, Aditi Kumar, Subhasish Tapadar, Adegboyega K. Oyelere, Smrruthi Vaidegi Venugopal, Shafiq A. Khan. Synthesis and biological evaluation of small molecules inhibitors targeting heterotrimeric Gαi2 protein [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 4596.
Abstract 46: 3D reconstruction and quantitative analysis of histology for prostate cancer
Authors: Ruusuvuori Pekka , Kartasalo Kimmo , Valkonen Mira , Valkonen Masi , Visakorpi Tapio , Nykter Matti , Latonen Leena
Published: August 2019

The integration of serial sectioning of tissue, digital whole slide imaging (WSI) and computational reconstruction algorithms enable the examination of histological samples in 3D at subcellular resolution. This allows visualizing and analyzing the imaged sample in its true three-dimensional context, offering a more comprehensive view on its spatial and morphological characteristics than that obtained via typical 2D examination. The advantages of reconstructing 3D models computationally using WSI over direct 3D imaging include the combination of high resolution, large sample sizes and compatibility with existing biochemical techniques such as in situ hybridization, immunohistochemistry and established histological staining and interpretation protocols.

For this purpose, we developed a software pipeline to perform routine 3D reconstruction tasks for large image sizes and datasets in a fully automatic manner. The steps in our 3D reconstruction process are image acquisition, alignment of images to a shared coordinate space, and visualization of the reconstructed 3D data. We optimized algorithm selection and computationally intensive hyperparameter tuning using a quantitative benchmarking framework and Bayesian optimization. Further, we applied our existing pipeline for feature based analysis of tissue and extended it into 3D, allowing quantification and visualization of hundreds of features characterizing the tissue.

We currently apply this system to characterize prostate cancer tumors. Prostate cancer is a heterogenous, often multifocal disease. In order to understand why and how certain tumor foci develop to a life-threatening disease over others, the tissue growth patterns and evolution of tumors with different genetic backgrounds need to be studied. We use mouse models of prostate cancer to study early tumor development connected to common genetic cancer alterations. By computing hundreds of features from the histology, and studying these in the spatial context, we gain important information of tumor characteristics as well as intra- and intertumor heterogeneity. In the future, we will further scale up the protocol to perform 3D reconstruction for serially sectioned human prostates in the near future.

Citation Format: Pekka Ruusuvuori, Kimmo Kartasalo, Mira Valkonen, Masi Valkonen, Tapio Visakorpi, Matti Nykter, Leena Latonen. 3D reconstruction and quantitative analysis of histology for 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 46.
Abstract 4602: Abi1 promotes breast cancer progression by regulating cell-matrix adhesion
Authors: Li Xiang , Regua Angelina , Bratslavsky Gennady , Kuznetsov Vladimir A. , Kotula Leszek
Published: August 2019

Breast cancer is one of the most common diagnosed cancers in the United States. There will be approximately 266,120 new breast cancer cases in the U.S. and over 40,000 patients are predicted to die from the malignant progression of their breast cancer tumors in 2018. Traditional therapeutic methods of the invasive breast cancer include surgery accompanied by chemotherapy. However, following the initial therapy patients often relapse and eventually die from tumor metastasis. Abl interactor 1, or Abi1, is an adaptor protein which regulates actin polymerization through participation in WAVE (Wiscott-Aldrich syndrome protein family verprolin homologous) complex. The WAVE regulatory complex (also called WRC), regulates actin based cellular behavior, such as cellular adhesion and cell migration, by activating Arp2/3 complex and promoting actin polymerization. Previous studies indicated the direct interaction between integrin α4 and Abi1, which suggests a potential role of Abi1 in regulating cell-matrix adhesion signaling by involved in the inside-out integrin signaling pathways. In order to test the above hypothesis, we developed two genetic engineered Abi1 knocked-out breast cell lines from either HER2+ breast cancer cell line SKBR3 or normal breast epithelial cell line MCF10A by using CRISPR-Cas9 technique. After confirming inactivation of Abi1 gene by Western blotting, DNA and RNA sequencing, cells were harvested and analyzed for integrins levels. Western blotting results indicated dysregulation of multiple integrins in the Abi1 knock-out cell lines. The results from subsequent immunofluorescence experiments of the Abi1 knock-out cell lines also suggested changes in cell-matrix adhesion signaling using staining for scaffold protein paxillin and phosphorylated Focal-Adhesion-Kinase (FAK) as readouts. Our findings suggest that Abi1 might functions as a key regulator in breast cancer tumor progression by playing an important role in regulating signal pathways in cell-matrix adhesion. Supported by the Carol Baldwin Foundation of CNY, and in part by NIH-NCI grant No. R01 CA161018.

Citation Format: Xiang Li, Angelina Regua, Gennady Bratslavsky, Vladimir A. Kuznetsov, Leszek Kotula. Abi1 promotes breast cancer progression by regulating cell-matrix adhesion [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 4602.
Abstract 4607: Androgen signaling is essential for prostate cancer development initiated from prostatic basal cells
Authors: He Yongfeng , Hooker Erika , Yu Eun-Jeong , Cunha Gerald R. , Liao Lan , Xu Jianming , Earl Andrew , Wu Huiqing , Gonzalgo Michael L. , Sun Zijie
Published: August 2019

Emerging evidence has shown that both prostatic basal and luminal cells are able to initiate oncogenic transformation. However, despite the diversity of tumor initiating cells, most prostate cancer cells express the androgen receptor (AR) and depend on androgens for their growth and expansion, implicating an essential role of androgen signaling in prostate tumorigenesis. Prostatic basal cells express p63 and are able to differentiate into luminal, neuroendocrine, and basal cells. In this study, we used a variety of relevant mouse models and in vivo systems to directly address the significance of androgen signaling in oncogenic transformation and tumor development initiated from prostatic p63-expressing cells. We demonstrate that activating Wnt oncogenic signaling by expressing stabilized b-catenin in prostatic p63-expressing cells is able to induce cell proliferation and the formation of atypical cell clusters in different prostatic lobes at embryonic, prepubescent, and adult stages. Intriguingly, despite the androgen insensitive nature of prostatic p63-expressing cells, androgens are still essential for these cells to grow and develop to androgen-dependent, luminal cell type prostate tumors. These findings are consistent with what have been observed in human prostate cancers, in which the majority of tumor cells are androgen-sensitive and possess luminal cell properties, providing new insight into the molecular mechanisms for prostate cancer initiation and progression.

Citation Format: Yongfeng He, Erika Hooker, Eun-Jeong Yu, Gerald R. Cunha, Lan Liao, Jianming Xu, Andrew Earl, Huiqing Wu, Michael L. Gonzalgo, Zijie Sun. Androgen signaling is essential for prostate cancer development initiated from prostatic basal cells [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 4607.
Abstract 4620: Development of orthotopic breast cancer brain metastasis patient-derived xenograft(PDX) model
Authors: Oshi Masanori
Published: August 2019

Background: The vast majority of the mortality of breast cancer results from distant metastasis and pre-clinical models are in critical need for effective drug development. Patient-derived xenograft (PDX) maintains the features of the donor tumors such as intra-tumor heterogeneity; however, PDX model of breast cancer brain metastasis that delivers stable consistent results is not popularized. We demonstrate our novel methods (coined “Oshi”method) used to develop an orthotopic brain metastasis patient-derived xenograft model (PDMOX) for breast cancer brain metastasis via tumor tissue implantation. Methods: Brain metastasis PDX were created using metastatic brain tumors from breast cancer patients and implanting them in the brain of NSG female mice. Tumors of ~1 mm3 were implanted mechanically minced tissue with medium. We implant the tissue through a frontal bone burr hole into the right caudate putamen. Tumor growth was monitored by magnetic resonance imaging (MRI). Results: “Oshi” method utilizes a pipette tip in order to inoculate tumor at the same depth. A minced tumor was instilled using a 23G needle as the “non-oshi” method. One hour post-surgical survival after implantation of minced tumor by “non-oshi” method was only 37.5% (3/8), whereas 100% (40/40) by the “Oshi” method. All tumors were well engrafted in surviving mice after both methods. There was larger variation in tumor growth after “non-oshi” (Median 20±25.0 day, range: 12-24 day. Tumor volume: median 5.6±21.0 mm3, range 2.8-48.7 mm3). One mouse developed ptosis, and 2 out of 3 mice that underwent the “non-oshi” method had sudden death. On the other hand, all mice that underwent the “Oshi” method lived until the tumor grew to 125-200 mm3 without neurological symptoms. The engraftment rate of Mincing tumor were 100% at 1.5 months after implantation, while the engraftment rate of Enzymatic digestion tumors was only 50% at 2 months. Mincing tumor had faster growth speed than the tumor of enzymatic digestion. The engraftment time of enzymatic digestion tumor was slower than other tumor tissue forms. All 3rd generation tumors could be detected at the point of 1month after implantation, on the other hand, it took 2 month to detect 1st generation tumors. The growth of 3rd generation tumors was significantly faster and bigger than others. Conclusion: Various surgical techniques used to generate PDMOX breast cancer models showed major differences in the tumors and outcomes. These novel models are expected to become powerful tools for preclinical studies in metastatic breast cancer.

Citation Format: Masanori Oshi. Development of orthotopic breast cancer brain metastasis patient-derived xenograft(PDX) model [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 4620.
Abstract 4630: In vivo characterization of the Duffy antigen receptor for chemokines (DARC/ACKR1) in breast cancer tumor progression
Authors: Martini Rachel N. , Jenkins Brittany D. , Newman Lisa A. , Manley Nancy , Davis Melissa B.
Published: August 2019

In studies of the tumor microenvironment (TME), factors that influence immune cell infiltration are of great interest, as these populations can influence disease prognosis, and potential treatment for patients. Through in silico analysis of TCGA breast cancer (BC) cohort data, we have identified the Duffy antigen receptor for chemokine/atypical chemokine receptor 1 (DARC/ACKR1) as a potential driver of immune cell infiltration in the BC TME. DARC/ACKR1 is an atypical chemokine receptor that modulates levels of chemokine both in circulation through expression on red blood cells, and participates in chemokine transcytosis in tissues through expression on both post-capillary venules of endothelial cells and epithelial cells. As chemokines establish gradients to recruit specific immune cell types, we hypothesize that the function of DARC/ACKR1 in chemokine level modulation also influences tumor-associated immune cell levels. In the TCGA BC cohort, we observe a strongly positive and significant correlation between DARC/ACKR1 expression and total number of tumor-associated leukocytes, specifically populations of B cells, T cell, monocytes and macrophages. To further characterize the role of DARC/ACKR1 in vivo, we have established a novel DARC knock-out BC transgenic mouse model to study DARC/ACKR1 in the BC TME. To develop our target experimental mice, Ackr1-/- female mice were crossed with Ackr1 +/-; C3(1)Tag +/0 to generate the target Ackr1 -/-; C3(1)Tag +/0 experimental mice. Experimental mice and the littermate controls were aged to 3.5 months, at which point they were euthanized. Blood was collected from the mice to characterize the circulating chemokine profile. Mammary glands containing tumors were fixed and paraffin embedded, followed by IHC and immunofluorescent staining for expression of DARC/ACKR1, target chemokines, immune cells, epithelial cells and endothelial cells. The remaining glands were dissociated, followed by fluorescent-activating cell sorting analysis (FACS) to quantify abundance of immune cells in the gland. Using these methods, we have defined the circulating chemokine profile alongside patterns of immune cell infiltration in our target DARC/ACKR1 knockout BC mice compared to littermate controls in mouse mammary tumors.

Citation Format: Rachel N. Martini, Brittany D. Jenkins, Lisa A. Newman, Nancy Manley, Melissa B. Davis. In vivo characterization of the Duffy antigen receptor for chemokines (DARC/ACKR1) in breast cancer tumor progression [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 4630.
Abstract 4631: Modeling the vascular sarcoma spectrum with genetically engineered mice
Authors: Hanna Jason A. , Langdon Casey G. , Garcia Matthew R. , Finkelstein David , Rehg Jerold E. , Hatley Mark E.
Published: August 2019

Angiosarcomas are highly aggressive vascular sarcomas with an extremely poor prognosis. Angiosarcomas can develop spontaneously or are associated with prior radiation, chronic lymphedema, or exposure to toxic chemicals such as vinyl chloride. Sequencing efforts have identified a number of genetic alterations in angiosarcoma, however the genetic drivers and actionable targets remain unclear. Despite the poor outcome for patients, there are limited resources for studying angiosarcoma, highlighting the need for genetic, in vivo models to better elucidate the underlying biology of the disease. In studying the role of DICER1 and microRNAs in mouse models of tumorigenesis, we found that Dicer1 deletion with aP2-Cre leads to aggressive and metastatic angiosarcoma development with 100% penetrance. ERK and S6 hyperphosphorylation in the aP2-Cre;Dicer1cKO (AD) tumors suggest Dicer1 loss results in activation of the RAS-MEK-ERK and mTOR pathways. To determine if direct activation of the these pathways could similarly transform aP2-Cre expressing cells we first interrogated the combination of oncogenic KrasG12D with Cdkn2a inactivation. We found that aP2-Cre;LSL-KrasG12D;Cdkn2acKO (AKC) mice rapidly develop angiosarcomas providing an accelerated model for assessing cooperating alleles and therapies. To activate the mTOR pathway we tested conditional Tsc1 deletion with aP2-Cre and found that all aP2-Cre;Tsc1cKO (AT) animals develop vascular tumors in the paws. In contrast to the AD and AKC tumors, AT tumors express markers of lymphatic endothelial cells such as PROX1. In addition, the tumors display a distinct nodular spindle cell-like morphology that is consistent with a low grade angiosarcoma resembling human kaposiform hemangioendotheliomas, a vascular tumor occurring predominantly in the extremities of infants. The distinct onset, growth kinetics, anatomic locations, and histologic presentation of the tumors from AD, AT, and AKC mice provide a mechanism to interrogate the drivers of angiosarcoma in less aggressive AT paw tumors and more aggressive AD and AKC tumors. Furthermore, the aP2-Cre driven mouse models provide a platform to study angiosarcoma initiation, progression, and metastasis for the identification of novel therapeutics to improve outcomes for this understudied and devastating disease.

Citation Format: Jason A. Hanna, Casey G. Langdon, Matthew R. Garcia, David Finkelstein, Jerold E. Rehg, Mark E. Hatley. Modeling the vascular sarcoma spectrum with genetically engineered mice [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 4631.
Abstract 4653: Melanocyte specific deletion of Map3k1 reveals its role in BRAFV600E-driven melanoma
Authors: Trucco Lucas D. , Mundra Piyushkumar A. , Garcia-Martinez Pablo , Hogan Kate , Dhomen Nathalie , Pavet Valeria , Marais Richard
Published: August 2019

BRAF is the most common driver oncogene in cutaneous melanoma. Activating mutations in the BRAF gene are enriched in melanomas associated with intermittent sun exposure, with the most frequent mutations occurring at codon 600, leading to a constitutive activation of the RAF/MEK/ERK signaling pathway. In previous studies, we demonstrated that ultraviolet radiation (UVR) accelerates melanomagenesis in a mouse model of melanoma driven by BRAFV600E. From whole exome sequencing analysis of the tumors we found that Map3k1 was the most recurrently mutated gene across different experimental cohorts. MAP3K1 is unique in having both a kinase and E3 ligase function, allowing it to regulate protein phosphorylation and ubiquitin-mediated proteasome degradation. The vast majority of the mutations in Map3k1 occurred within the RING domain, which contains the E2 binding site for ubiquitin-conjugating enzymes. The most prevalent non-synonymous mutations where present in codon 484 and arose as a consequence of C-to-T transitions at a dipyrimidine site, characteristic of UVR-induced DNA damage. To understand better the role of MAP3K1, we generated Tyr::CreERT2+/o;Map3k1f/f mice allowing us to conditionally delete Map3k1 specifically in the melanocytes upon topical application of tamoxifen to the dorsal skin of juvenile mice. These mice did not develop tumors, but when crossed to mice carrying a Braf+/LSL-V600E allele, all mice develop melanoma, with a shorter latency and an increased multiplicity compared to Tyr::CreERT2+/o;Braf+/LSL-V600E mice. Tumors showed similar histopathological features, but melanomas carrying Map3k1 floxed or mutant variants presented higher levels of phosphorylated ERK1/2 than those with wild-type Map3k1. In addition, analysis of expression data of human cutaneous melanomas from The Cancer Genome Atlas showed an inverse correlation between the expression of MAP3K1 and ERK2 protein levels. Taken together, our results suggest that MAP3K1 loss-of-function cooperates with BRAFV600E to drive melanoma development by increasing MEK-ERK pathway activation.

Citation Format: Lucas D. Trucco, Piyushkumar A. Mundra, Pablo Garcia-Martinez, Kate Hogan, Nathalie Dhomen, Valeria Pavet, Richard Marais. Melanocyte specific deletion of Map3k1 reveals its role in BRAFV600E-driven melanoma [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 4653.
Abstract 4656: TRAF1 is critical for regulating the BRAF/MEK/ERK pathway in non-small cell lung carcinogenesis
Authors: Wang Qiushi , Zhang Tianshun , Gao Ge , Bode Ann , Dong Zigang
Published: August 2019

Tumor necrosis factor receptor (TNFR)-associated factor 1 (TRAF1) is a unique TRAF protein that can interact directly or indirectly with multiple TNFR family members, regulatory proteins, kinases, and adaptors that contribute to its diverse functions in specific tissues. However, the role of TRAF1 in non-small cell lung cancer (NSCLC) remains unknown. In this study, we report that TRAF1 is overexpressed in human lung cancer cells and tissues. TRAF1 expression level inversely correlated with patient survival probability. Loss of TRAF1 decelerated tumor invasion in a urethane-induced lung carcinogenesis mouse model. Furthermore, TRAF1 expression affected TRAF2-mediated BRAF Lys48-linked ubiquitination, which was followed by the inhibition of growth and differentiation, and the induction of death in lung cancer cells. Overall, our work suggests that TRAF1 plays a novel role in the regulation of the BRAF/MEK/ERK signaling pathway in NSCLC and offers a candidate molecular target for lung cancer prevention and therapy.

Citation Format: Qiushi Wang, Tianshun Zhang, Ge Gao, Ann Bode, Zigang Dong. TRAF1 is critical for regulating the BRAF/MEK/ERK pathway in non-small cell lung carcinogenesis [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 4656.
Abstract 4657: MB21D2 amplification and recurrent mutation exhibit oncogenic activity through centrosome and cilia alteration
Authors: Gracilla Daniel Esguerra , Korla Praveen Kumar , Lai Louis , Sheu Jim Jinn-Chyuan
Published: August 2019

MB21D2 (Mab-21 containing domain 2), a member of the Mab family, is predicted to form protein complexes with different partners, thus performs diverse functions in cells. In silico analyses using TCGA data revealed MB21D2 amplification/overexpression in various types of human cancer and correlated with poor survivals in patients with head and neck and endometrial cancer. More interestingly, a recurrent Q311E mutation was found in the Mab-21 domain critical for protein-protein interactions and downstream signaling. When expressed in oral cancer TW206 and CAL27 cells (low endogenous MB21D2), wild-type MB21D2 moderately increased cell proliferation, migration, and invasion as compared to the vector control. Significantly, the mutant variant (Q311E) showed stronger oncogenic effects on those processes and formed bigger spheres in a 3-D culture condition. Data mining, functional network predictions, and Western blotting confirmed the involvement of MB21D2 in centrosome-associated ciliogenesis via a crosstalk between PIK3CA and PKA (GSK and CREB) signaling pathways. Blockage of such interactome by specific inhibitors against PIK3CA or PKA reduced oncogenic activities mediated by MB21D2. Our data therefore suggest MB21D2 amplification/overexpression or the Q311E mutation as cancer drivers promoting cancer development through alteration of centrosome and primary cilia activity.

Citation Format: Daniel Esguerra Gracilla, Praveen Kumar Korla, Louis Lai, Jim Jinn-Chyuan Sheu. MB21D2 amplification and recurrent mutation exhibit oncogenic activity through centrosome and cilia alteration [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 4657.
Abstract 4664: Massively parallel sequencing-based analysis of synchronous gastric cancer and lung cancer
Authors: Wu Nandie , Li Rutian , Shao Yang , Wang Lifeng , Liu Baorui , Wei Jia
Published: August 2019

Synchronous gastric cancer(GC) and lung cancer(LC) have been suggested to represent independent primary tumors rather than metastatic disease. We subjected sporadic synchronous GC/LC from five patients to whole-exome massively parallel sequencing, which revealed a median of and 1174 and 830 nonsynonymous somatic mutations in the synchronous GC and LC repectively. DNA mutations range between 292 to 2421 in lung samples and 287 to 1995 in stomach sample. Common mutations varies between 41 and 145. The substitution of C>A, C>T and T>C accounts for the most common mutations in both lung and stomach cancers. Tumor mutation burden (TMB ) varies from 2.7 to 11.0 Mut/Mb in Lung cancer and from 1.8 to 30.4 Mut/Mb in gastric cancer. We further did pathway analysis based on the list of mutant genes. Top 20 pathways enriched in each patient were analyzed. We found MAPK signaling pathway was most commonly enriched pathways in lung cancer patients, whereas PI3K-Akt signaling and Hedgehog signaling pathway were most commonly enriched pathways in stomach cancer patients. This result suggests the potential benefit of targeted therapeutic treatments involved in these pathways.

Citation Format: Nandie Wu, Rutian Li, Yang Shao, Lifeng Wang, Baorui Liu, Jia Wei. Massively parallel sequencing-based analysis of synchronous gastric cancer and 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 4664.
Abstract 4666: p21CIP1 promotes mammary cancer initiating cells via activation of Wnt/TCF1/CyclinD1 signaling
Authors: Hazan Rachel B.
Published: August 2019

Cancer stem cells (CSCs) generate and sustain tumors due to tumor initiating potential, resulting in recurrence or metastasis. We showed previously that knockout of the cell cycle inhibitor, p21CIP1, in the PyMT mammary tumor model, inhibits metastasis; however the exact mechanism remained unknown. Here, we show a pivotal role for p21 in potentiating a cancer stem cell-like phenotype. p21 knockout in PyMT mammary tumor cells suppressed tumorsphere formation, ALDH1 expression, and tumor initiating potential, that were in turn rescued by p21 overexpression. Consistent with these effects, p21 loss caused mesenchymal to epithelial transition that was accompanied by downregulation of Slug, Sox9, basal keratins and upregulation of luminal keratins and Claudin-3. Furthermore, p21 knockout suppressed Wnt/β-catenin signaling which was due to TCF1 downregulation. In turn, TCF1 knockdown in PyMT cells, attenuated Wnt signaling and tumorsphere formation. Interestingly, p21 rescue in p21 knockout cells, caused dramatic upregulation of Cyclin D1, consistent with known inhibition of Cyclin D1 nuclear export by p21. These data suggest that p21 promotes a cancer stem like phenotype via Wnt-TCF1 signaling along with direct upregulation of Cyclin D1 by p21.

Citation Format: Rachel B. Hazan. p21CIP1 promotes mammary cancer initiating cells via activation of Wnt/TCF1/CyclinD1 signaling [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 4666.
Abstract 4672: WDR26 regulates an AKT-Gsk3-Wnt/b-catenin signaling cascade to maintain the breast cancer stem cell population and controls cancer metastasis
Authors: Bhargava Dharmendra K. , Wang Wei , Lensing Maddison , Chen Songhai
Published: August 2019

Cancer metastasis is the major cause of tumor mortality and has been attributed in part to the presence of a minority subpopulation of cancer stem cells (CSCs) in the bulk of tumor cells. We showed previously that WDR26, a scaffolding/adaptor protein that is highly upregulated in breast cancer, promotes breast cancer growth and metastasis. Here we show WDR26 was required for maintaining the CSC populations in breast cancer cells and the formation of lung metastases. Downregulation of WDR26 in breast cancer cells impaired the CSC-like activities and reduced the CSC population. Mammary gland-specific deletion of WDR26 in the MMTV-PyMT mouse model of breast cancer had a little effect on primary tumor formation but largely abolished spontaneous lung metastasis. WDR26 promoted β-catenin activation via AKT and GSK3, and the activity of AKT, GSK3 and β-catenin was required for maintaining the CSC population in breast cancer cells. Our results have identified a novel, WDR26-dependent pathway that links breast CSC activities to tumor metastatic potential.

Citation Format: Dharmendra K. Bhargava, Wei Wang, Maddison Lensing, Songhai Chen. WDR26 regulates an AKT-Gsk3-Wnt/b-catenin signaling cascade to maintain the breast cancer stem cell population and controls cancer 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 4672.