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  • Undergraduate Poster Abstracts
  • Cancer Biology

    FRI-934 EXPLORING THE MICROBIOME OF CERVICAL CANCER THROUGH TRANSKINGDOM NETWORKS

    • Khiem Lam ;
    • Jialu Hu ;
    • Andrey Morgun ;

    FRI-934

    EXPLORING THE MICROBIOME OF CERVICAL CANCER THROUGH TRANSKINGDOM NETWORKS

    Khiem Lam, Jialu Hu, Andrey Morgun.

    Oregon State University, Corvallis, OR.

    Cervical cancer is the second most common cancer in women worldwide. Recently, the role of microbiota in some types of cancer has been shown. Therefore, to understand if microbes contribute in any way to the development and progress of cervical cancer, we investigated the cervical microbiome via analysis of 16S rRNA sequencing data. Using qPCR, we detected bacterial DNA in 118 out of 133 samples of cervical carcinoma. However, 16s library preparation and sequencing was possible for only 58 samples with higher amounts of bacterial DNA. After matching our results with the Human Microbiome Project, we observed that cancer samples compositionally represent a very tight cluster, with its closest neighbors in principle coordinate analysis being samples from the vagina, stool, and skin. Overall, the alpha diversity of cervical cancer communities was quite low (Shannon index 3.88) compared to the healthy microbiome of other body sites. Furthermore, the most abundant taxa in these samples were of phyla Bacteroidetes (39%), Firmicutes (25%), Fusobacteria (14%), and Proteobacteria (9%). To get a better understanding of the structure of cancer bacterial communities, we have reconstructed a microbial covariation network. This work presents the previously unexplored microbiome of invasive cervical cancer. Despite its low diversity, the cervical cancer microbiome is a complex structure not limited to a few microbes. The analysis of transkingdom interactions is under way to reveal the place of bacteria in tumor, host, and HPV interactions for the development of cervical cancer.

    THU-945 RE-SENSITIZATION TO DOXORUBICIN-INDUCED CYTOTOXICITY IN DRUG-RESISTANT BREAST CANCER CELLS BY GREEN TEA POLYPHENOL

    • Clarissa Hernandez ;
    • Kamaleshwar Singh ;
    • Logeswari Ponnusamy ;

    THU-945

    RE-SENSITIZATION TO DOXORUBICIN-INDUCED CYTOTOXICITY IN DRUG-RESISTANT BREAST CANCER CELLS BY GREEN TEA POLYPHENOL

    Clarissa Hernandez, Kamaleshwar Singh, Logeswari Ponnusamy.

    Texas Tech University, Lubbock, TX.

    Breast cancer is the second most prevalent cancer-related cause of death in women. Despite advances in chemotherapeutic treatments, acquisition of resistance in the course of treatment causes failure in successful clinical management. Drug resistance is multifactorial, and accumulating evidence implicates aberrant expression of genes leading to increased survival as one of the mechanisms for drug-resistant cancer cells. Recent studies show that epigallocatechin gallate-3 (EGCG), a natural polyphenol derived from green tea, may potentially inhibit cancer cell growth and has no toxicity to healthy cells. However, the effect of EGCG in restoring the chemotherapeutic drug-induced cytotoxicity in drug-resistant breast cancer cells is not known. Therefore, the objective of this study was to evaluate the effect of EGCG on doxorubicin-resistant MCF-7 breast cancer cells. Doxorubicin-sensitive and resistant MCF-7 cell lines were either first pre-treated with EGCG (25 µM final concentration) for 24 hours prior to doxorubicin (100 nM final concentration) treatment, or directly treated with doxorubicin for 48 hours. The doxorubicin-induced cytotoxicity was measured by MTT assay. The results of MTT assay indicate a 20% increase in cell death in doxorubicin resistant cells that were first pretreated with EGCG followed by doxorubicin than the cells that were treated with doxorubicin without EGCG pre-treatment. Results of this study suggest that pretreatment with EGCG could potentially enhance sensitivity to doxorubicin-induced cytotoxicity in doxorubicin-resistant MCF-7 breast cancer cells. Further study of cell cycle analysis and gene expression changes would be helpful in understanding the molecular mechanism for EGCG-mediated re-sensitization to chemotherapy in drug-resistant breast cancer cells.

    THU-923 EFFECTS OF IN VIVO KNOCKDOWN OF A COLD-INDUCIBLE RNA-BINDING PROTEIN (CIRP) ON BREAST TUMORIGENESIS

    • Joey Ochoa ;
    • Daniel Lujan ;
    • Rebecca S. Hartley ;
    • Helen Hathaway ;

    THU-923

    EFFECTS OF IN VIVO KNOCKDOWN OF A COLD-INDUCIBLE RNA-BINDING PROTEIN (CIRP) ON BREAST TUMORIGENESIS

    Joey Ochoa, Daniel Lujan, Rebecca S. Hartley, Helen Hathaway.

    The University of New Mexico, Albuquerque, NM.

    RNA-binding proteins (RBPs) are essential to processes that involve RNAs, beginning from their transcription through to their degradation. In this study, we utilized the polyomavirus middle T antigen (PyMT) mouse model to evaluate the role of cold-inducible RBP (CIRP) in breast cancer. When PyMT is expressed in the mammary epithelium of mice, it causes aggressive tumorigenesis. CIRP has been shown to incite the translation of mRNAs encoding stress-induced proteins and is observed to have increased expression in breast cancer. We have shown that human CIRP overexpression in the PyMT mouse appears to inhibit breast tumorigenesis during early stages. Based on these results, the current study set out to evaluate the effects of CIRP knockdown on tumor development. We expect knockdown to allow greater progression of tumor growth, as well as a higher rate of metastasis. To address this hypothesis, CIRP will be knocked down using siRNA in a PyMT -mammary tumor- derived cell line. PyMT cells transiently transfected with siCIRP and siControl will be injected into the mammary fat pad of a wild-type mouse, and tumor growth will be assessed. CIRP knockdown in PyMT cells is being analyzed via RT-qPCR and western blotting to determine the effective timing for mammary fat pad injections. Tumor growth will be monitored every 3 days for 3 to 6 weeks. Final tumor burden and metastases will be assessed as will tumor progression, proliferation, and apoptosis. The results of this study will tell us if CIRP overexpression has the potential to protect against breast cancer.

    FRI-932 R-KETOROLAC HAS A NEGATIVE EFFECT ON OVARIAN CANCER METASTASIS, PROLIFERATION, AND MIGRATION

    • Jordan Weisend ;
    • Angela Wandinger-Ness ;
    • Elsa Romero ;
    • Yuna Guo ;

    FRI-932

    R-KETOROLAC HAS A NEGATIVE EFFECT ON OVARIAN CANCER METASTASIS, PROLIFERATION, AND MIGRATION

    Jordan Weisend, Angela Wandinger-Ness, Elsa Romero, Yuna Guo.

    The University of New Mexico, Albuquerque, Albuquerque, NM.

    Being that ovarian cancer has few to no symptoms in its early stages, late detection is very common, making the disease the fifth most deadly cancer in women. Our group has identified a GTPase targeted drug that has been shown to negatively affect metastasis, proliferation, and migration in ovarian cancer cells. We have used monolayer representations of the ovarian tumors for numerous experiments, but scientists have recently determined that these traditional cultures are unrealistic in representing the in vivo environment in which tumor cells thrive. Here, we implement a method of 3D culture that better represents the peritoneal cavity in which ovarian cancer develops and spreads. Using fibroblast and mesothelial cells embedded in matrix proteins, we are able to closely represent the extracellular matrix and the internal organ linings through which tumor cells must migrate to metastasize to other sites. Subsequent addition of fluorescently labeled ovarian cancer cells allows us to quantify the number of cells able to invade and migrate through the membrane. Based on previous research, it has been shown that the R enantiomer of ketorolac has a negative effect on the migration of ovarian tumor cells while S-ketorolac is inactive. Testing both drugs alongside a control, showed R-ketorolac significantly inhibits 3D invasion. Using these results, we are able to confirm that the samples containing R-ketorolac show a decreased amount of invading ovarian cancer cells. These results pave the way for future research both on stem cells and GTPase regulation of the metastasis processes in ovarian cancer.

    FRI-937 DNA LIGASE IV MODULATES THE CELLULAR RESPONSE TO DNA REPLICATION STRESS

    • Melissa Joe ;
    • Amanda Ashley ;

    FRI-937

    DNA LIGASE IV MODULATES THE CELLULAR RESPONSE TO DNA REPLICATION STRESS

    Melissa Joe, Amanda Ashley.

    New Mexico State University, Las Cruces, NM.

    Correctly repairing DNA damage is crucial to the survival and genomic integrity of cells. Double strand breaks (DSBs) are particularly problematic DNA damage; failure to properly repair DSBs can precede genomic instability leading to cancer. Paradoxically, because of their high proliferative index, replication toxins are successfully used to treat cancer. The 2 major DNA DSB repair pathways are homologous recombination and nonhomologous end-joining (NHEJ). DNA ligase IV is a central component of NHEJ, performing the final ligation step to reseal the broken DNA ends in concert with XRCC4. We determined cells lacking another NHEJ protein, DNA-PK, restarted DNA replication forks quicker than wildtype following replication stress, though whether other NHEJ proteins are involved is unclear. Our goal was to investigate whether ligase IV, a protein whose cellular functions are exclusive to NHEJ, is involved in the replication stress response. Our hypothesis is ligase IV delays replication restart following exposure to replication toxins. To test our hypothesis, we performed cell culture, qPCR, western-blot analysis, EdU incorporation, CellTiter-Glo viability assay, and DNA fiber assay. We confirmed our ligase IV knockdown via qPCR and western blotting. Cells had low viability when exposed to siRNA against ligase IV compared to non-target siRNA. A higher percentage of cells exposed to non-targeting siRNA incorporated EdU compared to ligase IV knockdown. Replication stress is a major driver of chromosomal instability and development of drug resistance. Understanding how NHEJ proteins may impact the replication stress response may provide opportunities for developing more effective cancer treatments.

    FRI-945 ANALYSIS OF PRC2 PROTEIN-PROTEIN INTERACTIONS AND EZH2 INHIBITION WITH SMALL MOLECULES IN B-CELL LYMPHOMA CELL LINES

    • Lindsay Redman ;
    • Anneke Kramm ;
    • Stuart Schreiber ;

    FRI-945

    ANALYSIS OF PRC2 PROTEIN-PROTEIN INTERACTIONS AND EZH2 INHIBITION WITH SMALL MOLECULES IN B-CELL LYMPHOMA CELL LINES

    Lindsay Redman1, Anneke Kramm2, Stuart Schreiber2.

    1New Mexico State University, Las Cruces, NM, 2Center for the Science of Therapeutics, The Broad Institute of MIT & Harvard, Cambridge, MA.

    Chromatin compaction, largely influenced by posttranslational histone modifications, is one of many points of regulation for gene expression. The enhancer of zeste homolog 2 (EZH2) subunit of the human polycomb repressive complex 2 (PRC2) catalyzes trimethylation of the histone H3 lysine 27 (H3K27me3), which is important for recruiting regulatory factors involved in the repression of transcription. Overexpression of, or activating mutations in EZH2 have been implicated in many cancers, due to silencing of tumor suppressor genes via high levels of H3K27me3. The use of small molecule inhibitors to target many histone methyltransferase (HMT) enzymes such as EZH2 has become an increasingly attractive area of therapeutic development. Here we attempt to use cellular thermal shift assays (CETSA) to study these protein-compound interactions as well as testing the efficacy of EZH2 inhibitors on H3K27 methylation levels in several B-cell lymphoma cell lines. Through CETSA, we have shown the stabilization of several PRC2 complex proteins by the binding of GSK126, an EZH2 small molecule inhibitor. In addition, we show inhibition/decrease of H3K27me3 in both EZH2 wild-type and mutant lymphoma cell lines with GSK126, with the mutant having a greater sensitivity to compound treatment. In the exploration and further optimization of the novel technique CETSA, there is great potential for development of high throughput assays assessing small molecule inhibition and eventually leading to pharmaceutical development of small molecule inhibitors.

    FRI-926 DEFINING THE ROLE OF A MUTANT ISOCITRATE DEHYDROGENASE (IDH) IN MALIGNANT GLIOMAS

    • Joseph-Michael Fields ;
    • Gemma Robinson ;
    • Beatrice Philip ;
    • Sheri Holmen ;

    FRI-926

    DEFINING THE ROLE OF A MUTANT ISOCITRATE DEHYDROGENASE (IDH) IN MALIGNANT GLIOMAS

    Joseph-Michael Fields1, Gemma Robinson2, Beatrice Philip2, Sheri Holmen2.

    1Langston University, Langston, OK, 2The University of Utah, Salt Lake City, UT.

    Gliomas are the most common primary central nervous system tumors, but the molecular mechanisms responsible for the development and progression of these tumors are far from being completely understood. Mutations in the metabolic enzyme isocitrate dehydrogenase (IDH) were recently found in approximately 80% of WHO grade II-III gliomas and secondary glioblastomas. These mutations inhibit the enzyme's ability to convert isocitrate to a-ketoglutarate and, instead, confer a novel gain-of-function resulting in the conversion of a-ketoglutarate to 2-hydroxyglutarate. However, the fundamental mechanism(s) by which these mutations affect glioma cell growth remain unclear. This study aims to further our understanding of the function of mutant IDH using an established brain-tumor mouse model. Together, these approaches will lead to a better understanding of the biology of mutant IDH gliomas and will help guide the development of new therapies to improve survival and reduce morbidity in these patients.

    FRI-935 TARGETING BREAST CANCER VIA VIRAL CHEMOTHERAPY

    • Rigoberto Perez Hernandez ;
    • Jorge G. Gomez-Gutierrez ;
    • Heshan Sam Zhou ;
    • Rajesh Sharma ;
    • Kelly M. McMasters ;

    FRI-935

    TARGETING BREAST CANCER VIA VIRAL CHEMOTHERAPY

    Rigoberto Perez Hernandez1, , Jorge G. Gomez-Gutierrez2, Heshan Sam Zhou2, Rajesh Sharma2, Kelly M. McMasters2.

    1Division of Nutritional Sciences, Cornell University, Ithaca, NY, 2University of Louisville School of Medicine, Louisville, KY.

    Tamoxifen (TAM) resistance is a major clinical challenge in the treatment of breast cancer (BC). The resistance mechanism of TAM-treated BC cells is pro-survival autophagy. Autophagy is the basic mechanism that involves cell degradation of unnecessary or dysfunctional cellular components. On the other hand, oncolytic adenoviruses (OAds) selectively kill cancer cells by viral oncolysis leaving normal cells intact. The purpose of this study was to evaluate the capacity of TAM-induced autophagy in enhancing OAd-mediated oncolysis in human and murine BC cells. Estrogen receptor positive (ER+) BC (human MCF-7 and murine 4T1) and estrogen receptor negative (ER-) BC (MDA-MB-231) cell lines were infected with an OAd (E1B-deleted Adhz60) alone or in combination with TAM or temozolomide (TMZ). Previously, our group showed that TMZ increased virotherapy potency in other cancer cells. A crystal violet staining assay revealed that the combination therapy of Adhz60 with TAM or TMZ resulted in greater cell killing (< 30% cell viability) compared to single therapy and controls (p < 0.05). The Adhz60-mediated oncolysis increased in the presence of TAM only in MCF-7 cells. Interestingly, TMZ-enhanced virotherapy potency in all 3 cell lines. Western blot showed that enhanced oncolysis was associated with an increase of adenoviral E1A expression. To the best of our knowledge, this is the first time that TAM has been used to increase ER+ BC cell virotherapy. More importantly, we also discovered that TMZ enhanced Adhz60-mediated oncolysis independently of ER status. Our results provide evidence of a novel combinatorial regimen as an alternative therapy to treat BC.

    THU-859 DETERMINING THE ROLE OF FAK AND PYK2 IN PREVENTING DNA DAMAGE SIGNALS

    • Ramilyn Gonzales ;
    • Isabelle Tancioni ;
    • David Schlaepfer ;

    THU-859

    DETERMINING THE ROLE OF FAK AND PYK2 IN PREVENTING DNA DAMAGE SIGNALS

    Ramilyn Gonzales1, Isabelle Tancioni2, David Schlaepfer2.

    1Chaminade University of Honolulu, Honolulu, HI, 2University of California, San Diego, La Jolla, CA.

    Focal adhesion kinase (FAK) and proline-rich tyrosine (Pyk2) are proteins that regulate endothelial cell survival, proliferation, and motility. Endothelial cells are a luminal tissue layer in blood vessels and function to regulate the flow of nutrients from the blood to surrounding tissue and to export catabolic waste. Previous results from Schlaepfer Lab showed that FAK induces degradation of p53, a tumor-suppressor transcription factor that leads damaged cells to apoptosis. This experiment is investigating the in vivo endothelial effects of DNA damage signals in mice lungs when FAK and Pyk2 are knocked out. Genetically engineered mice models were raised with Pyk2 knocked out (Pyk2 null). The Pyk2 null mice have induced FAK manipulations in endothelial cells only: wild-type (WT), kinase-dead (KD), and double knocked-out (DKO). Previous results showed that elimination of FAK and Pyk2 causes endothelial cell dysfunction as demonstrated by the presence of perivascular edema in the lung tissue. We analyzed the expression of phospho-histone H2AX, a DNA damage repair signal, in lung sections by immunofluorescence confocal microscopy. Preliminary results showed an increase in phospho-histone H2AX expression in DKO mice. Future investigations can be conducted on endothelial cell death via p53 cell cycle regulation through its association with p21.

    THU-933 THE G PROTEIN-COUPLED ESTROGEN RECEPTOR PROMOTES TUMORIGENESIS THROUGH TUMOR-ASSOCIATED FIBROBLASTS

    • Rachael Couch ;
    • Helen Hathaway ;

    THU-933

    THE G PROTEIN-COUPLED ESTROGEN RECEPTOR PROMOTES TUMORIGENESIS THROUGH TUMOR-ASSOCIATED FIBROBLASTS

    Rachael Couch1, Helen Hathaway2.

    1The University of Texas at Dallas, Richardson, TX, 2The University of New Mexico, Albuquerque, NM.

    The steroid and estrogen sex hormone 17β- estradiol (E2) is required for normal development of the female mammary gland but also promotes proliferation in breast cancer. E2 acts by binding to 3 known estrogen receptors. Breast-tumor expression of the most recently discovered receptor, the G protein-coupled estrogen receptor (GPER), has been clinically associated with increased metastasis. Additionally, GPER is expressed by and promotes proliferation of breast cancer-associated fibroblasts (CAFs) in the tumor microenvironment. Since CAFs can also promote tumor progression and increased metastasis, it is unclear if GPER actions in tumor cells, CAFs, or both, could be pro-tumorigenic. In our study, we aimed to define the interaction between E2 and the tumor microenvironment in tumor progression via GPER mediation by examining mice tumors treated with GPER agonists or antagonists for the presence of altered extracellular matrix (ECM), a product of CAFs. Additionally, 3D culture models of mouse breast cancer cells treated with similar compounds, cultured with or without mammary fibroblasts, were examined for GPER-dependent activation of fibroblasts, proliferation, and increase in motile phenotype. Our studies show that GPER activates mammary fibroblasts in vitro to create tumorigenic CAFs, providing tumor stroma GPER expression as a possible therapeutic target. Paradoxically, our results seem to suggest that, in vivo, GPER reduces collagen I fibrils, and moreover tumor cell proliferation appeared to be minimally affected by GPER inhibition. Future investigations will determine if collagen expression or fibrillogenesis is altered in tumors. Finally, tumor cell migration and invasion in the presence of GPER modulation may determine if these pro-tumorigenic behaviors contribute to GPER-dependent metastasis.

    THU-858 TRIPTERINE DERIVATIVES AGAINST DEXAMETHASONE-RESISTANT ACUTE LYMPHOBLASTIC LEUKEMIA

    • Kara Anderson ;
    • Fatima Rivas ;
    • Taotao Ling ;

    THU-858

    TRIPTERINE DERIVATIVES AGAINST DEXAMETHASONE-RESISTANT ACUTE LYMPHOBLASTIC LEUKEMIA

    Kara Anderson1, Fatima Rivas2, Taotao Ling2.

    1Marshall University, Huntington, WV, 2St. Jude Children's Research Hospital, Memphis, TN.

    Acute lymphoblastic leukemia (ALL) is one of the most common pediatric cancers in developed countries. Although ALL has a high cure rate (> 94%), up to a quarter of patients relapse and, at that stage, their prognosis is poor. Refractory ALL cases are commonly associated with drug resistance, particularly to glucocorticoid (GC) treatment, an integral component of the therapy. In a previous companion study, certain natural product (NP) compounds were identified as potential therapeutic agents based on their ability to preferentially inhibit proliferation in GC resistant ALL stable cell lines Nalm06-R and UOCB-1R. Elucidation of the structure of these compounds has prompted further investigation of one NP: a triterpenoid isolated from the ethonolic extraction of Tripterygium wilfordii. The triterpenoid family of NPs has demonstrated anticancer properties in solid tumors but has not been extensively studied in hematological cancers. The high bioactivity of this NP in our leukemic model led to the evaluation of its chemical constituents. The main components and the synthesized derivatives of the main component displayed a promising EC50 in the single-digit micromolar range against GC-resistant ALL cells and showed a therapeutic window in control cell lines. Our mechanistic studies also indicate that S-phase cell cycle arrest is occurring in the treated cell lines with apoptosis being mediated through activation of caspase-3/7. Herein, we highlight our promising findings concerning this triterpenoid-based NP molecular scaffold against GC resistant ALL.

    FRI-848 INVESTIGATING THE SPECIFICITY OF BMI1 INHIBITION BY SMALL MOLECULE PTC-209 IN LUNG AND SKIN CANCER IN VITRO MODELS

    • Timothy Musoke ;
    • Rachit Neupane ;
    • Jacqueline Lees ;

    FRI-848

    INVESTIGATING THE SPECIFICITY OF BMI1 INHIBITION BY SMALL MOLECULE PTC-209 IN LUNG AND SKIN CANCER IN VITRO MODELS

    Timothy Musoke1, Rachit Neupane2, Jacqueline Lees2.

    1University of Massachusetts Boston, Boston, MA, 2Massachusetts Institute of Technology, Cambridge, MA.

    Bmi1is a component of the polycomb repressive complex (PRC1) which maintains stable repression of loci by ubiquitylating lysine 119 in histone H2A. One of the established targets of the PRC1 is the p16INK4A/p19Arf locus, which encodes for the important p16 and p19 tumor suppressors that regulate cell proliferation and senescence. Bmi1 is overexpressed in multiple cancer types and has a well-established role as an oncogene and stemness regulator; hence, it is a great candidate for anticancer therapy. Previous studies have identified a small-molecule compound, PTC-209, as a potent Bmi1 inhibitor using human colorectal cancer cell culture and patient-derived xenograft (pdx) models. In order to explore the specificity of Bmi1 inhibition, we used 2 in vitro models: genetically isogenic paired non-small-cell lung cancer murine cell lines that are either Bmi1 wildtype or null, and genetically isogenic-paired human melanoma cell lines that have either normal or a knocked down level of Bmi1. In both models, crystal violet staining was used to assess proliferation on drug treatment, and biochemical analysis using the western blot assay was used to assay Bmi1 level. Small molecule inhibition of Bmi1 is a powerful tool to study the biology of Bmi1 and a new avenue for cancer therapy. Understanding the specificity of PTC-209 will elucidate whether it is a viable candidate to pursue in that regard.

    FRI-936 IDENTIFYING THE POST-TRANSLATIONAL REGULATORY MECHANISMS IMPACTING DIFFERENTIAL CELF1 STABILITY IN THE EPITHELIAL TO MESENCHYMAL TRANSITION

    • Shebna Cheema ;
    • Arindam Chaudhury ;
    • Julia Chernis ;
    • Joel Neilson ;

    FRI-936

    IDENTIFYING THE POST-TRANSLATIONAL REGULATORY MECHANISMS IMPACTING DIFFERENTIAL CELF1 STABILITY IN THE EPITHELIAL TO MESENCHYMAL TRANSITION

    Shebna Cheema1, Arindam Chaudhury2, Julia Chernis3, Joel Neilson2.

    1University of Houston-Downtown, Houston, TX, 2Baylor College of Medicine, Houston, TX, 3The University of Texas at Austin, Austin, TX.

    The Neilson laboratory has identified the CUGBP and embryonically lethal factor 1(CELF1) RNA binding protein as a novel driver of epithelial to mesenchymal transition (EMT) in breast epithelial cells. Similar to what has been observed in cellular models of myotonic dystrophy, CELF1’s stability and function in MCF-10A cells are correlated with phosphorylation of the protein on serine (S) and threonine (T) residues by specific isoforms of protein kinase C (PKC). However, the specific S/T residues within CELF1 that are phosphorylated by PKC remain unidentified. To identify these phosphorylation sites for downstream functional studies, we have undertaken 2 parallel approaches. Computational analysis using the KinasePhos2.0 webtool revealed 25 S/T residues predicted to be phosphorylated by PKC in CELF1. Using gene synthesis, we mutated each of these predicted sites to alanine in CELF1 coding sequence. Each of the 25 predicted PKC sites was then restored individually via site-directed mutagenesis to generate a battery of constructs containing single putative phosphorylation sites. Each of these constructs is being subcloned into a glutathione S transferase (GST) gene fusion system for affinity purification and in vitro kinase assays. We have also immunoprecipitated GFP-CELF1 fusion proteins from mesenchymal MCF10A cells treated either with a PKC inhibitor or vehicle control, and submitted gel-purified protein bands for mass spectrometry. Between these complementary approaches, we hope to directly identify S/T residues that are phosphorylated by PKC during EMT. This will facilitate future studies to identify if and how such modification influences CELF1 protein degradation.

    FRI-933 TEMPOL IN PRIMATE PREGNANCY AND FETAL ANTIRETROVIRAL DRUG GENOTOXICITY

    • Ana Serrato ;
    • Ofelia Olivero ;

    FRI-933

    TEMPOL IN PRIMATE PREGNANCY AND FETAL ANTIRETROVIRAL DRUG GENOTOXICITY

    Ana Serrato, Ofelia Olivero.

    National Cancer Institute, Bethesda, MD.

    Antiretroviral (ARV) drug combinations used as therapy for HIV-1/AIDS are given during pregnancy and for 6 wks after birth to eliminate maternal-fetal HIV-1 transmission. To study ARV drug-induced effects in pregnancy, we used Erythrocebus patas (patas) monkey dams exposed to human-equivalent protocols containing the ARV drugs zidovudine (AZT) and lamivudine (3TC). The AZT/3TC combination causes bone marrow cell genotoxicity in newborn patas infants exposed in utero. Here, we used Tempol, a cytoprotective antioxidant, along with AZT/3TC, for the last half of gestation, hypothesizing that Tempol would protect the infants against the AZT/3TC-induced genotoxicity. Bone marrow cells were cultured from patas infants at birth, and approximately1,000 cells per treatment group were examined for the presence of micronuclei (MN), a genotoxic end point. Exposure groups included Tempol (n = 3), Tempol/AZT/3TC (n = 1), and control (n = 1). Historical AZT/3TC positive controls gave values of 23 MN/1,000 cells, while unexposed controls showed 11 MN/1,000 cells. Tempol/AZT/3TC exposed cells only had 20 MN/1,000 cells. Therefore, Tempol did protect the patas against the genotoxic end point, MN. This patas study has been designed to model human clinical protocols for HIV treatment in pregnancy and models potential outcomes we may see in humans.

    THU-935 ROLE OF NUCLEAR ABL TYROSINE KINASE IN CISPLATIN-INDUCED APOPTOSIS

    • Thalia Jimenez ;
    • Priya Sridevi ;
    • Jean Wang ;

    THU-935

    ROLE OF NUCLEAR ABL TYROSINE KINASE IN CISPLATIN-INDUCED APOPTOSIS

    Thalia Jimenez, Priya Sridevi, Jean Wang.

    University of California, San Diego, La Jolla, CA.

    Cisplatin is a DNA-damaging drug that is used to treat many cancers, but it also causes acute kidney injury. In response to DNA damage, a complex network of pathways is activated to coordinate DNA repair with cell proliferation and cell death. Double-stranded breaks generated by cisplatin-activate nuclear Abl tyrosine kinase, which phosphorylates a host of nuclear proteins to regulate the responses. Renal proximal tubule epithelial cells (RPTC) are sensitive to cisplatin and undergo DNA-damage-induced apoptosis. It is well established that p53 activates the expression of PUMA to stimulate apoptosis. The goal of this project is to test the hypothesis that Abl stimulates PUMA protein expression to cause apoptosis. Recombinant PUMA plasmids will be transfected into cells with wild-type (WT) or nuclear import-defective (NID) Abl. Cells will be treated with cisplatin, PUMA RNA will be measured by quantitative real-time PCR, and PUMA protein will be measured by western blotting. We expect that cisplatin will activate Abl in the WT, but not the NID cells. Furthermore, we expect that cisplatin will stimulate PUMA protein in the WT, but not the NID cells. This study is the first to investigate the translational regulation of PUMA protein. Basic mechanisms elucidated from this study will provide clues on how to inhibit PUMA protein expression in RPTC to preserve these cells. Furthermore, knowledge gained from this research will provide clues on how to stimulate PUMA protein expression in cancer cells as a therapeutic strategy to kill cancer cells.

    THU-944 THE CYTOTOXIC EFFECTS OF CROCIN ON H1650 LUNG CANCER CELLS

    • Erika Ramos ;
    • Meghan Cromie ;
    • Weimin Gao ;

    THU-944

    THE CYTOTOXIC EFFECTS OF CROCIN ON H1650 LUNG CANCER CELLS

    Erika Ramos, Meghan Cromie, Weimin Gao.

    Texas Tech University, Lubbock, TX.

    Lung cancer is the primary cause of cancer-related deaths in both men and women in the U.S. More individuals die of lung cancer than colon, breast, and prostate cancers combined. Patients diagnosed with lung cancer often experience a variety of side effects during conventional treatment and eventually may develop resistance. Therefore, alternative therapeutics with fewer or no side effects are needed. Crocin, natural carotenoid compound found in crocus and gardenia flowers, has exhibited therapeutic potential in cancer cell lines based with limited data. The purpose of this study is to determine the effects of crocin on H1650 lung cancer cells using MTT assay, quantitative reverse-transcriptase PCR (qRT-PCR), and western blot to evaluate cytotoxicity, gene expression, and protein expression, respectively. A dose-dependent increase in cytotoxicity was observed after H1650 cells were treated with crocin, with an observed IC50 of 23 mg/mL for 24 h and 18 mg/mL for 48 h. No changes in the gene expressions of EGFR or survivin were observed. However, p21 demonstrated a 2.5-fold upregulation after 48 h treatment with 16 mg/mL crocin, which was further confirmed by western blot. P21 is a well-known inhibitor of cell proliferation through cell cycle arrest. Thus, the significant upregulation of p21 suggests that crocin could inhibit lung cancer cell growth. The findings of this study warrant further examination into the usefulness of crocin for lung cancer treatment and its molecular mechanisms.

    THU-936 CONTRIBUTIONS AND MECHANISMS OF FOXO TRANSCRIPTION FACTORS ON THE PI3K PATHWAY

    • Andrea Salinas ;
    • Megan Keniry ;
    • Itzel Flores ;
    • Lilia Sanchez ;
    • Cristina Rodriguez ;
    • Neftali Vazquez ;
    • Rebecca Marks ;
    • Jesse Hirschmann ;

    THU-936

    CONTRIBUTIONS AND MECHANISMS OF FOXO TRANSCRIPTION FACTORS ON THE PI3K PATHWAY

    Andrea Salinas, Megan Keniry, Itzel Flores, Lilia Sanchez, Cristina Rodriguez, Neftali Vazquez, Rebecca Marks, Jesse Hirschmann.

    University of Texas Pan-American, Edinburg, TX.

    The conserved phosphatidylinositol 3 kinase (PI3K pathway) impacts cellular growth, metabolism, and survival. PI3K is a lipid kinase that catalyzes the production of phosphatidylinositol 3,4,5 trisphosphate (PIP3), which in turn binds to and activates targets such as the protein kinase AKT. Over 20 substrates have been characterized for AKT; arguably the best studied AKT substrates are the Forkhead subfamily O (FOXO) transcription factors: FOXO1, FOXO3, and FOXO4. AKT phosphorylates FOXO1, 3, and 4 on conserved amino acids, leading to their cytoplasmic retention/inactivation. Canonically, FOXO transcription factors induce genes that promote cell cycle arrest and apoptosis. However, one or more of these factors promotes PI3K output via a homeostatic feedback mechanism. The purpose of this poster is to addresses the mechanisms, contributions, and differences between FOXO1, 3, and 4. FOXO1 on PI3K pathway signals transduction. Currently, we are taking loss of function and gain of function approaches to delineate the specific contributions of each FOXO factor on the PI3K output (in mammalian cell lines, mouse embryonic fibroblast, and established human cell lines). We are utilizing western blot analysis and quantitative reverse transcript PCR to examine PI3K output. Excitingly, we are finding that FOXO factors have unique contributions to PI3K.

    THU-937 THE ASSOCIATION OF SEX-STRATIFIED MGMT DNA REPAIR SNPS IN MELANOMA RISK

    • Christopher Hughes ;
    • Jenna Lilyquist ;
    • Kirsten White ;
    • Salina Torres ;
    • Li Luo ;
    • DeAnn Lazovich ;
    • Marianne Berwick ;

    THU-937

    THE ASSOCIATION OF SEX-STRATIFIED MGMT DNA REPAIR SNPS IN MELANOMA RISK

    Christopher Hughes1, Jenna Lilyquist1, Kirsten White1, Salina Torres1, Li Luo1, DeAnn Lazovich2, Marianne Berwick1.

    1The University of New Mexico, Albuquerque, NM, 2University of Minnesota, Minneapolis, MN.

    Cutaneous melanoma utilizes a complex and dynamic route to pathogenesis that is not entirely understood. Incidence rates over the past several decades have risen and fueled investigations to elucidate the causes of melanoma. Melanoma is the 6th most common cancer in the U.S. with an estimated 73,870 new cases and 9,940 deaths predicted in 2015. Males experience higher mortality and incidence rates than females, but little is known about the sex-dependent biological differences for risk. Genetic factors, including DNA repair genes, have been reported to play a role in melanomagenesis. The O6-methylguanine-DNA methyltransferase (MGMT) gene encodes an enzyme involved in DNA repair, and single nucleotide polymorphisms (SNPs) in this gene are implicated in melanoma risk and survival. We genotyped 893 melanoma cases and 766 control participants from the Minnesota Skin Health Study for 24 SNPs in MGMT. Males had an increase in risk for rs4750766 (p = 0.04, OR 1.45, 95% CI = 1.14 - 1.83) and a decrease in risk for rs7075505 (p = 0.002, OR 0.71, 95% CI = 0.05 - 0.98). There were no significant variants associated with melanoma risk in females. When controlling for multiple tests, the SNPs did not meet correction for false discovery; however, DNA repair variants in MGMT may still play a role in sex-dependent risk for melanoma. Although MGMT’s exact influence on melanomagenesis remains unknown, further investigation into the function of these SNPs could provide useful insight into sex-specific risk.

    THU-934 DEFINING THE ROLE OF MACROPHAGES IN HEAD AND NECK CANCER

    • Hilda Mejia Pena ;
    • Judith Varner ;

    THU-934

    DEFINING THE ROLE OF MACROPHAGES IN HEAD AND NECK CANCER

    Hilda Mejia Pena1, Judith Varner2.

    1San Diego City College, San Diego, CA, 2University of California, San Diego, La Jolla, CA.

    Macrophages are bone-marrow-derived phagocytic cells that accumulate in tissues responding to infections and damage. These cells can be polarized into 2 distinct states that either promote M1 and/or M2 inflammation. While M1 macrophages promote inflammation secondary to chronic infections that may predispose to cancer, M2 macrophages promote tumor growth by suppressing T-cell mediated anti-tumor immunity. Head and neck squamous cell carcinomas are aggressive tumors that can exhibit extensive infiltration by immunosuppressive macrophages. We will test the hypothesis that head and neck tumor-secreted factors induce immune suppressive M2 macrophage polarization in vitro and in vivo. We also hypothesize that inhibitors of M2 polarization can prevent head and neck tumor growth by preventing immune suppression. To accomplish these aims, we will test the effects of head and neck carcinoma-secreted cytokines on macrophage polarization in vitro by incubating primary macrophages with tumor cell-conditioned medium and evaluating the macrophage gene expression profiles in response to these factors. We will also characterize the evolution of the immune response to head and neck tumor cell inoculation in vivo in syngeneic mice by characterizing the invasion of tumors by natural killer cells, T cells, B cells, and M1 and M2 macrophages. Finally, we will evaluate the effects of altering macrophage polarity on the growth of these tumors in vivo. Because head and neck tumor cells may induce immunosuppressive macrophage polarization, drugs that alter macrophage polarity might be effective in combination with other conventional strategies to overcome immunological tolerance and promote tumor regression.

    FRI-944 THE EFFECTS OF A PHOTO ISOMERIZING COMPOUND TREATMENT ON P-GLYCOPROTEIN DRUG-RESISTANT CANCER

    • Justin Avila ;
    • Aryana Cruz ;
    • Zachary Jordan ;
    • Steve Holliday ;
    • Nuha Kadri ;
    • Matthew Gdovin ;

    FRI-944

    THE EFFECTS OF A PHOTO ISOMERIZING COMPOUND TREATMENT ON P-GLYCOPROTEIN DRUG-RESISTANT CANCER

    Justin Avila1, Aryana Cruz2, Zachary Jordan1, Steve Holliday1, Nuha Kadri1, Matthew Gdovin1.

    1The University of Texas at San Antonio, San Antonio, TX, 3The University of Texas Health Science Center at San Antonio, San Antonio, TX.

    Chemotherapeutics are one of the most commonly employed methods for the treatment of cancers. However, their use inadvertently selects for the survival of cancer cells resistant to these drugs. The over expression of P-glycoprotein, a non-specific active transporter in cancers, is the most common mechanism of multidrug-resistance (MDR). The appearance of MDR cancer often leads to a terminal prognosis in patients since there are limited treatment options. Recent studies in our laboratory have shown that a photo isomerizing compound (PIC) is able to passively diffuse into several cancer cell lines. The PIC is then activated with light, causing the release of hydrogen ions, reducing the intracellular pH (pHi) and apoptotic cell death in vitro. Due to the properties of the PIC and mode of activation, we hypothesize that the PIC acidification treatment will be able to induce apoptosis in a MDR cancer line that employs P-glycoprotein as its mechanism of drug-resistance. An MDR prostate cancer cell line 22-RV-1, which over expresses P-glycoprotein, will be loaded with PIC and the pH-sensitive fluorophore DCFDA to monitor acidification. After light activation of the PIC, cells will be exposed to Annexin V and Ethidium Homodimer III to monitor for apoptosis and necrosis, respectively. Based on its size, we predict that the PIC will not be shuttled out of the cell by P-glycoproteins, and will be able to induce apoptosis upon activation. These findings will provide a foundation for the use of the PIC to treat drug resistant cancers. (Partially funded by GM060655.)

    FRI-858 IDENTIFYING NOVEL BIOMARKERS IN URINE FROM CANINE TRANSITONAL CELL CARCINOMA PATIENTS

    • George Pope ;
    • Shay Bracha ;

    FRI-858

    IDENTIFYING NOVEL BIOMARKERS IN URINE FROM CANINE TRANSITONAL CELL CARCINOMA PATIENTS

    George Pope, Shay Bracha.

    Oregon State University, Corvallis, OR.

    The purpose of this study is to identify biomarkers for canines with transitional cell carcinoma (TCC). The urine of canines with infected bladders, normal bladders, or TCC were collected and subjected to both lipid identification and exosome miRNA sequencing. A pilot study was done in which lipids were shown to be present in the urine samples through the use of the resin HP20 and tested against chloroform on thin-layer chromatography. Lipids were extracted through lyophilization and speed vac as described before, and subjected to mass spectrometry analysis. The differences between the lipids present in each category of urine will be determined, and potential biomarkers for TCC will be identified. Additionally, exosomes will be isolated from the same urine samples and analyzed by miRNA sequencing. The resulting sequence information will be compiled and analyzed to detect differences in miRNA expression between the patients with and without TCC. If the results of these tests show viable differences in the urine samples, then future extensions of this project will be to develop tests that can identify these red flags during simple urine sampling on routine visits to the veterinarian. Through cross-species analyses, we may also be able to show correlations between canine TCC and human bladder cancer, perhaps showing shared biomarkers and leading to improved tests for bladder cancer in humans as well.

    THU-848 MITOCHONDRIAL MASS AND FUNCTION IN BREAST CANCER: SIGNIFICANCE FOR CANCER SUBTYPING AND TREATMENT

    • Hector Galvez ;
    • Kay Macleod ;

    THU-848

    MITOCHONDRIAL MASS AND FUNCTION IN BREAST CANCER: SIGNIFICANCE FOR CANCER SUBTYPING AND TREATMENT

    Hector Galvez1, Kay Macleod2.

    1California State University, San Marcos, San Marcos, CA, 2Ben May Department for Cancer Research, University of Chicago, Chicago, IL.

    Prior studies have yet to address the role mitochondrial mass has on the viability of breast cancer cells. Cells rely on mitochondria for the production of ATP through glycolysis and oxidative phosphorylation. In this study, we aim to uncover the function that mitochondrial mass has in primary breast cancer cells, and whether it can serve as a future therapeutic target. A high-throughput screen was conducted to isolate FDA-approved drugs that modulated mitochondrial mass. Cell lines representing different molecular subtypes of breast cancer were cultured and treated with the top screen hit, mycophenolic acid (MPA), which was shown to increase mitochondrial mass. All cell lines showed increased mass after 48 hours as shown by change in fluorescence intensities of MitoTracker Green, a fluorophore that localizes to mitochondria regardless of membrane potential. Cell viability was analyzed using propidium iodide (PI) stain, which is able to penetrate the cell membrane of dead cells but not that of living cells. After 48 hr treatment MPA, all cells showed an increase in cell death as compared to untreated samples. Increases in superoxide along with sensitivity to MPA despite inhibition of biogenesis suggests MPA serves as an inhibitor to mitophagy. These findings suggest mitochondrial mass may serve as a future therapeutic target. Treatments will expand to cover primary breast cancers representing all molecular subtypes to better understand if certain subtypes have increased sensitivity to MPA. Future studies aim to uncover the precise metabolic mechanism responsible for the increase in mass.

    FRI-859 MOLECULAR DISSECTION OF WNT SIGNALING IN THE PCSD NEW SERIES OF PATIENT-DERIVED XENOGRAFT MODELS OF BONE METASTATIC PROSTATE CANCER

    • Theresa Mendoza ;
    • Danielle Burner ;
    • Michelle Muldong ;
    • Amy Strasner ;
    • Christina Wu ;
    • Seung Chol Park ;
    • Elana Godebu ;
    • Omer Raheem ;
    • Catriona Jamieson ;
    • Colm Morissey ;
    • Isaac Kim ;
    • Anna Kulidjian ;
    • Christina Jamieson ;

    FRI-859

    MOLECULAR DISSECTION OF WNT SIGNALING IN THE PCSD NEW SERIES OF PATIENT-DERIVED XENOGRAFT MODELS OF BONE METASTATIC PROSTATE CANCER

    Theresa Mendoza1, Danielle Burner1, Michelle Muldong1, Amy Strasner1, Christina Wu1, Seung Chol Park1, Elana Godebu1, Omer Raheem1, Catriona Jamieson1, Colm Morissey2, Isaac Kim3, Anna Kulidjian1, Christina Jamieson1.

    1University of California, San Diego, La Jolla,, CA, 2University of Washington, Seattle, WA, 3Rutgers Cancer Institute of New Jersey, New Brunswick, NJ.

    One of the leading health problems affecting men in the U.S. and many other parts of the world is prostate cancer. Cancer patients are surviving longer due to androgen deprivation therapies. However, over 80% of advanced patients develop bone metastatic prostate cancer for which there is no cure. Therefore, the goal of this project is to determine which signaling components would serve as potential targets for bone metastatic prostate cancer therapy using our lab’s new prostate cancer xenograft models. Researchers have shown that the WNT signaling pathway regulates various processes that are important for cancer progression including tumor initiation, tumor growth, cell death, differentiation, and metastasis. Thus, it is hypothesized that the WNT signaling componentsWNT5A, BMP-6, and ROR receptors help prostate cancer thrive and become aggressive in the bone microenvironment in the absence of androgen. To determine whether these WNT signaling components were expressed at the RNA level in our lab’s prostate cancer xenograft models, reverse transcriptase polymerase chain reactions (RT PCR) have been performed. The results of the experiments showed that BMP-6, ROR1, and WNT5A components are expressed in our prostate cancer xenograft models. Further testing using real time PCR (qPCR) and immunohistostaining assays will be done in the future and, if the results of these experiments indicate that the WNT signaling pathway could potentially serve as an effective target for therapy, we would then utilize knock-out and overexpression methods on our prostate cancer xenograft models.

    THU-925 THE EFFECTS OF SORAFENIB ON HEAD AND NECK CANCER

    • Clayton Speed ;
    • John Robert Hatherill ;

    THU-925

    THE EFFECTS OF SORAFENIB ON HEAD AND NECK CANCER

    Clayton Speed, John Robert Hatherill.

    Del Mar College, Corpus Christi, TX.

    Head and neck squamous cell carcinoma (HNSCC) is the 6th most malignant cancer. Standard treatment is combined surgery, chemotherapy, and radiation; even with treatment, 60% of patients do not live more than 5 years after diagnosis. New hope for HNCSS patients has come in the form of Sorfenib, a novel kinase inhibitor which is FDA-approved to treat kidney cancers. Previous studies have shown that Sorfenib, which was designed to inhibit mutant B-Raf oncoprotein in renal carcinoma, radiosensitizes HNSCC cell lines and retards proliferation. Interestingly, HNSCC cell lines are B-Raf wildtype, meaning that Sorafenib must be inhibiting secondary kinases, causing the overserved phenotypes in HNSCC lines. We hypothesize that the polo-like kinase (PLK1) is the secondary target of Sorafenib, causing the observed radiosensitivity and DNA repair defects. PLK1 does not directly act in any of these processes but phosphorylates the enzyme nucleophosmin (NPM), a poly-modified protein that has major roles in DNA repair and cell cycle regulation. It is also highly expressed in rapidly proliferating cells. NPM’s role in proliferation and DNA repair is controlled by phosphorylation via PLK1. Western blot and mass spectroscopy were used to map global protein modification before and after combined Sorfenib and X-ray treatment. We found that phosphorylation of NPM happens within minutes after irradiation, but cells treated with Sorfenib never activate NPM. We conclude that because NPM was not phosphorylated in Sorfenib-treated cells after DNA damage, PLK1 must be a novel secondary target of Sorfenib and a promising candidate for targeted therapy in radioresistant tumors.

    FRI-925 INVESTIGATION OF THE PROCESSING OF THE EWS-FLI1 FUSION TRANSCRIPT

    • Guillermo Rangel Rivera ;
    • Natasha Caplen ;

    FRI-925

    INVESTIGATION OF THE PROCESSING OF THE EWS-FLI1 FUSION TRANSCRIPT

    Guillermo Rangel Rivera, Natasha Caplen.

    Center for Cancer Research, National Cancer Institute, Bethesda, MD.

    Ewing sarcomas (ES) are bone and soft tissue tumors affecting children. Most ES tumors (approximately 85%) harbor a translocation between chromosomes 22 and 11 [t(11:22) (q24;q12)] which results in the generation of the fusion oncogene EWS-FLI1. EWS-FLI1 codes for the oncoprotein EWS-FLI1, an aberrant transcription factor required for ES tumorigenesis. Recently, a genome-wide loss-of-function (LOF) RNAi screen identified several genes involved in RNA processing as required for EWS-FLI1 activity. Due to the complexity of EWS-FLI1 gene organization, we hypothesized that constitutive expression of specific RNA processing proteins are required for the expression of EWS-FLI1 and hence EWS-FLI1 activity. Understanding the mechanism by which RNA processing regulates EWS-FLI1 expression may reveal new druggable targets for ES. To this end, we validated genes from the primary RNAi screen using a combination of luciferase reporter assays, cell viability, and quantitative and qualitative PCR-based techniques in 4 different ES cell lines. Using a reporter assay of EWS-FLI1 activity and siRNA-mediated LOF, we identified XPA binding protein 2 (XAB2) as a lead candidate. In this preliminary study, silencing of XAB2 by RNAi leads to a 3.3-fold decrease in EWS-FLI1 mRNA and a decrease in EWS-FLI1 protein levels. ES cell viability decreased in a time-dependent manner by approximately 40 and 65%, 48 and 72 h, respectively, following XAB2 LOF. This data suggests silencing of XAB2 down regulates EWS-FLI1 expression, and thus decreases cell viability. However, the exact mechanism by which XAB2 regulates EWS-FLI1 expression is unknown and remains the subject of ongoing investigation.

    THU-924 GPER ACTIVATION AND ECM DEVELOPMENT IN METASTASIS

    • Andrew Gutierrez ;
    • Helen Hathaway ;
    • Nicole Marjon ;
    • Eric Prossnitz ;

    THU-924

    GPER ACTIVATION AND ECM DEVELOPMENT IN METASTASIS

    Andrew Gutierrez, Helen Hathaway, Nicole Marjon, Eric Prossnitz.

    The University of New Mexico, Albuquerque, NM.

    Breast cancer is ranked 2nd in cancer-related deaths among women. Estrogen receptors, primarily estrogen receptor α (ERα), promote breast cancer progression. G protein-coupled estrogen receptor (GPER) expression correlates with recurrence, reduced tumor size, and increased distant metastasis in vitro and in a breast cancer mouse model. However, little is known about the molecular mechanisms by which GPER promotes breast tumor metastasis; understanding GPER function in metastasis may lead to better breast cancer prevention and treatments. Increased extracellular matrix (ECM) protein production, notably collagen, promotes metastasis and tumor development. Since GPER expression correlates with tumor size and metastasis, we hypothesize that GPER promotes ECM microenvironment changes that support metastasis. To test this hypothesis, tissue sections from mammary glands of tumor-bearing mice treated with GPER activating (E2, G-1) and inhibiting (G36) ligands were stained with Masson’s trichrome to delineate the collagenous ECM. ECM was quantitated in 4 to 5 randomly selected fields per sample. Results demonstrate no statistically significant difference in any treatment groups compared to sham-treated mice. Our next goal is to examine expression of specific ECM proteins including collagen, laminin, and fibronectin using immunohistochemical methods. If our hypothesis is supported and ECM proteins are decreased when GPER is inhibited, we will be one step closer to understanding GPER function in breast cancer metastasis.

    THU-926 INTERACTIONS OF STRESS AND 3-IODOTHYRONAMINE TREATMENT ON GENE EXPRESSION IN MCF-7 CELLS

    • Christopher Ponce ;
    • Shaikh Rahman ;

    THU-926

    INTERACTIONS OF STRESS AND 3-IODOTHYRONAMINE TREATMENT ON GENE EXPRESSION IN MCF-7 CELLS

    Christopher Ponce, Shaikh Rahman.

    Texas Tech University, Lubbock, TX.

    The present study assessed the effects of 3-iodothyronamine (T1AM) treatment and cell-density mediated-oxidative stress on the expression of 3 different genes implicated in stress: hypoxia-inducible factor 1-alpha (HIF1α), pyruvate dehydrogenase alpha 1 (PDH1α), and lactate dehydrogenase alpha (LDHα). We hypothesized that both T1AM and cell-density mediated oxidative stress affect HIF1α, PDHα, and LDHα gene expression in MCF-7 cells. To test our hypothesis, non-invasive MCF-7 (ER+) breast cancer cells were cultured under standard culture conditions (5% CO2) and incubated at 37 °C. Cells were seeded at 50k, 100k, and 150k in Dulbeccos’s modified eagle’s medium (DMEM). The cells were then treated with either 50 mM T1AM or DMSO (control) for 24 hours. After that, cells were harvested for total RNA extraction and gene expression analyses. Cells in the plate those were seeded at 100k and treated with 50 mM T1AM showed moderate increase in HIF1α and LDHα gene expression. In conclusion, our data indicated that the cell-density mediated oxidative stress along with the T1AM treatment are responsible for moderate increase in H1F1α and LDHα gene expression in MCF-7 cells.

    FRI-924 TARGETING GROWTH AND INVASIVENESS IN CANCER CELLS

    • Stephanie Soriano ;
    • Nitika Parmar ;

    FRI-924

    TARGETING GROWTH AND INVASIVENESS IN CANCER CELLS

    Stephanie Soriano, Nitika Parmar.

    California State University Channel Islands, Camarillo, CA.

    Invasiveness is a characteristic feature of malignancy in cancer and can greatly reduce the 5-year survival rate of patients affected. This study aimed to determine the impact of 5 proposed anti-cancer agents (berberine chloride, azathioprine, gossypol, miltefosine, and etoposide) on 3 female cancer cell lines: MES-SA (uterine), C33A (cervical), and SKBR (breast). Invasiveness was tested using the CultreCoat® 96 Well Medium BME Cell Invasion Assay. The cancer cell lines were grown to 80 - 90% confluency and then treated with the proposed anti-cancer agents at 2 µM for 24 hours. Cells were harvested and subsequently seeded in the invasion chamber for 24 hours. Migration of the cells reflecting invasiveness was monitored via a fluorimetric assay and quantified. Untreated cells served as controls. Invasiveness was influenced by the type of reagent and cell line. Etoposide and berberine chloride greatly reduced invasiveness in a majority of the cell lines. A dose-dependent impact on invasiveness is currently being studied. Targeting invasiveness in these aggressive cell lines may lead to a potential direction.