bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 229: A cross-species approach to identifying genetic modifiers of BRCA1-associated breast cancer risk
Authors: Hu Hsien-Ming , Voytovich Uliana , Paulovich Amanda G.
Published: June 2012

Background: Although carriers of germline mutations of BRCA1 have a high risk of developing breast and ovarian cancers, penetrance is not complete, and it is likely that unidentified genes modify the risk for individual mutation carriers. We hypothesize that genes that modify the penetrance of cancer in BRCA1 carriers interact with BRCA1 mutations to confer increased sensitivity to DNA damage at the cellular level, and that cellular sensitivity to DNA damage can be used as an intermediate phenotype to predict breast cancer susceptibility in humans.

Objective: Our goal is to test a cross-species systems biology approach to predict gene-gene interactions underlying breast cancer susceptibility, using sensitivity to DNA damage as an intermediate phenotype.

Experiment Procedure: The yeast RAD9 gene is one putative homolog of the human breast tumor suppressor gene BRCA1. We conducted a genome-wide screen to identify genes that interact with a rad9 mutation to confer enhanced sensitivity to DNA damage. The yeast screen identified 25 interactions, from which we selected 10 candidate gene-gene interactions to be tested in human breast epithelial cells. To determine if the gene-gene interactions discovered in yeast are conserved in human mammary epithelial cells, we built stable human mammary epithelial cells (MCF10A) capable of single- or double-gene knockdown via lentivirus-mediated RNA interference. Cells confirmed for intended gene knockdown were subject to a clonogenic assay to determine if increased sensitivity to DNA damage can be observed in double gene vs. single gene knockdowns.

Results: Among the 10 pairs of interactions tested in MCF10A cells, two genes, HLTF and MGMT, were found to interact with BRCA1 to confer enhanced DNA damage sensitivity, based on a clonogenic assay. Cells with combined knockdown of BRCA1 and HLTF show increased sensitivity to a chronic low dose of MMS treatment compared with those with only single knockdown of BRCA1 or HLTF. In addition, distinct morphological features were observed in double knockdown cells with MMS treatment. Cells became enlarged and showed a flat, senescence-like morphology. Even a low concentration of 0.001% MMS induced a strong and irreversible growth arrest in cells with combined knockdown of BRCA1 and HLTF, but not in the single knockdown cells.

Conclusion: Our results suggest that it is feasible to use a cross-species (yeast-to-human) comparative systems genetics strategy to identify gene-gene and pathway-pathway interactions. Ongoing studies are aimed at determining whether any of the observed gene-gene interactions contribute to breast cancer penetrance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 229. doi:10.1158/1538-7445.AM2011-229
Abstract 2298: ck2-Dependent phosphorylation of progesterone receptors (PR) on Ser81 regulates PR-B-isoform-specific target gene expression in breast cancer cells
Authors: Hagan Christy R. , Regan Tarah , Dressing Gwen , Lange Carol
Published: June 2012

Progesterone receptors (PR) are critical mediators of mammary gland development and contribute to breast cancer progression. Progestin-induced rapid activation of cytoplasmic protein kinases leads to selective regulation of growth-promoting genes by phospho-PR species. Herein, we show that phosphorylation of PR Ser81 is ck2-dependent and progestin-regulated in intact cells, but also occurs in the absence of PR ligands, when cells enter the G1/S phase of the cell cycle. T47D breast cancer cells stably expressing a PR-B mutant that cannot be phosphorylated at Ser81 (S81A) formed fewer soft agar colonies. Regulation of selected genes by PR-B, but not PR-A, also required Ser81 phosphorylation for basal and/or progestin-regulated (BIRC3, HSD11β2, and HbEGF) expression. Additionally, wt PR-B, but not S81A mutant PR, was robustly recruited to a PRE-containing transcriptional enhancer region of BIRC3; abundant ck2 also associated with this region in cells expressing wt but not S81A PR. We conclude that phospho-Ser81 PR provides a platform for ck2 recruitment and regulation of selected PR-B target genes. Understanding how PR functions in the context of high kinase activities characteristic of breast cancer is critical to understanding the basis of tumor-specific changes in gene expression and will speed the development of highly selective treatments.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2298. doi:10.1158/1538-7445.AM2011-2298
Abstract 2320: In vivo characterization of small molecule inhibitors of replication protein A: Implications for cancer therapy
Authors: Neher Tracy M. , Bodenmiller Diane M. , Turchi John J. , Jalal Shadia I.
Published: June 2012

The nucleotide excision repair (NER) pathway catalyzes the removal of bulky DNA adducts from the genome and aids in maintaining genomic stability. The NER pathway has two subsets; global-genomic NER (GG-NER) and transcription coupled NER (TC-NER). They differ mainly in steps involving the recognition of the DNA lesion. The NER pathway plays a very important role in the recognition, removal and repair of intra-strand adducts induced by cisplatin; a commonly used chemotherapeutic agent. Previous studies have demonstrated that a decrease in NER activity enhances cancer sensitivity to cisplatin. The recognition of DNA damage involves a series of proteins including the xeroderma pigmentosum group A and C proteins (XPA and XPC) and Replication Protein A (RPA). RPA, a single-strand DNA binding protein, plays an essential role in both NER sub-pathways in addition to DNA replication. This makes RPA an ideal protein to target cancer therapy. Previous studies from our lab identified compounds which disrupt RPA's ssDNA binding activity in vitro. We sought to further characterize these RPA small molecule inhibitors (SMI) in cancer cell culture models. Using lung and ovarian cancer cell lines, viability curves, apoptosis assays, cell cycle analysis, signaling and transcription regulation were analyzed with single agent treatment and in combination with cisplatin. Results demonstrate potent cell killing in single agent studies and lengthening of the G1/ S-phase of the cell cycle. RPA SMIs also induce cell death through a classical apoptotic pathway. Additionally, data demonstrate a synergistic relationship with cisplatin with sequential treatment. Cell signaling analyses and DNA repair gene expression studies are ongoing to further determine the DNA damage response induced by these compounds. Overall these data demonstrate the therapeutic potential of SMI of RPA in treatment of cancer as single agents and in combination with cisplatin.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2320. doi:10.1158/1538-7445.AM2011-2320
Abstract 2327: Structural and kinetic characterization of crizotinib with wild-type and mutant anaplastic lymphoma kinase
Authors: McTigue Michele A. , Timofeevski Sergei L. , Liu Wei , Deng Ya-Li , Marrone Tami , Cui Jean , Brooun Alexei
Published: June 2012

Dysregulation of Anaplastic Lymphoma Kinase (ALK), primarily through gene translocations, has been shown to be involved in a variety of cancers. Crizotinib, an orally available small molecule inhibitor of the ALK tyrosine kinase has demonstrated marked efficacy in clinical trials of NSCLC patients harboring the EML4-ALK oncogenic gene rearrangement. Mutation of some residues within the ALK kinase domain have been reported to confer acquired or de novo resistance to crizotinib. To understand the binding of crizotinib to ALK and the mechanism of resistance of specific mutations we generated and kinetically characterized wild-type (WT) and mutant ALK kinase domains (KD). Additionally, ALK KD crystal structures were determined of the WT nonphosphorylated apoenzyme and complexes with crizotinib bound to WT and a L1196M gatekeeper mutation. No large protein conformational changes are necessary for crizotinib to bind to unliganded ALK. The interactions which crizotinib makes with ALK are similar to its binding to c-Met with the exception of notable differences in the position of the activation loop between ALK and c-Met. Mutation of the L1196 gatekeeper residue to methionine results in a ∼8-fold increase in catalytic efficiency of phosphorylation of an activation loop peptide and also more rapid enzyme auto-phosphorylation. In addition, inhibition of L1196M ALK by crizotinib was reduced ∼9-fold, compared to wild-type enzyme, from Ki determinations. The crystal structures show that L1196 or M1196 make direct contact with crizotinib. The diminished activity of crizotinib against L1196M ALK is therefore likely due to both higher intrinsic kinase activity and a subtle change in the ALK-crizotinib binding interactions.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2327. doi:10.1158/1538-7445.AM2011-2327
Abstract 233: Crosstalk of Her2 signaling and cell cycle control in the response to targeted therapeutics in breast cancer
Authors: Rivadeneira Dayana , Mercier Isabelle , Lisanti Michael P. , Witkiewicz Agnieska , Knudsen Erik S.
Published: June 2012

Previous studies have shown that breast cancer is associated with genetic and molecular modifications, such as aberrant amplification of oncogenes and/or inactivation of tumor suppressors. This heterogeneity translates into a wide range of breast cancer subtypes that exhibit different disease behaviors and therapeutic sensitivity. A common oncogenic activation used to classify subtypes of breast cancer is the overexpression of human epidermal growth factor receptor 2 (Her2). These highly aggressive tumors present distinct characteristics in disease progression and therapeutic response. One of the main complications with current therapies is therapeutic bypass and disease recurrence. Therefore, a better understanding of the mechanisms of therapy resistance and the development of alternative downstream interventions is of utmost significance.

There are several signaling cascades involved in tumor development that can be activated by the Her2 receptor tyrosine kinases. These signals ultimately converge on the cell cycle machinery, leading to the functional inactivation of RB to promote aberrant cell cycle progression and cell growth. In this context, RB pathway inactivation has been associated with aggressive tumor behavior and poor prognosis in breast cancer. In the current study, we explore the impact of RB inactivation in Her2 overexpressing disease in the response to Her2/EGFR antagonists. Furthermore, we assess the efficacy of reactivating the RB-pathway as a downstream therapeutic alternative in the treatment of Her2 overexpressing tumors and therapy-resistant disease in vitro and in vivo.

In vitro studies demonstrate that RB loss can modulate the response to a Her2/EGFR antagonist. Alternatively, results indicate that a novel therapeutic agent that specifically targets cyclin dependent kinase 4 and 6 (CDK4/6) activity dramatically and efficiently inhibits acini/cell growth. Furthermore, CDK4/6 inhibition prevents the formation of multiacinar structures characteristic of Her2 overexpression. Furthermore, a Her2 overexpressing mouse mammary tumor model was utilized to confirm our in vitro findings. These studies indicate that CDK4/6 inhibition was effective in suppressing tumor cell proliferation in vivo. Combined, these findings provide further insight into the crosstalk between Her2 signaling and cell cycle control in the context of therapeutic response, and introduce a novel mode of therapeutic intervention.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 233. doi:10.1158/1538-7445.AM2011-233
Abstract 2344: Distinct combinations of signaling pathways promote migration of MDA-MB-231 breast carcinoma cells in response to different stimuli
Authors: Harrison Susan M. , Chen Min , Knifley Teresa , O'Connor Kathleen L.
Published: June 2012

Cancer cell invasion and metastasis are predicated on the activation of signaling pathways that control dynamic organization of cytoskeletal architecture. The coordinate reorganization of an actin network pushing at the leading edge and an actin-myosin mediated retraction at the rear of the cell facilitates cell polarity to allow directed movement. Motility of the highly invasive breast carcinoma cell line MDA-MB-231 is promoted by various stimuli including lysophosphatidic acid (LPA), hepatocyte growth factor (HGF) and epidermal growth factor (EGF). The goal of this project is to delineate the signaling pathways mediating motility in response to each of these stimuli. Identifying common and conditional pathways mediating cell motility has important implications for therapeutic intervention of cancer cell invasion and metastasis. We conducted migration, invasion, and morphology assays in the presence of inhibitors for pathways known to contribute to the cytoskeletal reorganization required for cell migration. LPA, HGF and EGF were evaluated for their capacity to signal through the Rho family small GTPases, Rho and Rac, the MAP kinase pathway via MEK1/2 and myosin light chain kinase (MLCK) to influence cell migration and invasion. LPA is a recognized component of breast cancer progression and potent stimulator of Rho activity, which has been extensively implicated in the migration of carcinoma cells in a variety of cancers including breast. Surprisingly, LPA-stimulated migration of MDA-MB-231 cells was not affected by Rho inhibitors but was instead sensitive to Rac inhibition. In contrast, Rho was required for HGF and EGF-stimulated migration of MDA-MB-231 cells while Rac had no effect on these processes. Other common pathways mediating cell motility including MLCK are used by all three chemoattractants. MDA-MB-231 cells employed all of the pathways examined for invasion, potentially reflecting the complexity of navigating in a three dimensional environment. These observations suggest that while multiple signaling pathways contribute to the migratory potential of a cancer cell, not all pathways are operating under all conditions. Different context-specific pathways may supplement a core of common pathways to create a functional signal but the characteristics of the response will likely rely on the particular combination used by the cell.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2344. doi:10.1158/1538-7445.AM2011-2344
Abstract 2355: Full-length wild-type CD99 increases the migration and invasiveness of human malignant glioma cells
Authors: Seol Ho Jun , Nam Do-Hyun , Lee Yu Jin , Smith Christian A. , Rutka James T.
Published: June 2012

The malignant glioma is the most common primary human brain tumor, and its migration and invasiveness away from the primary tumor mass is considered a leading cause of tumor recurrence and treatment failure. Recently, gene expression profiling revealed CD99, which is a transmembrane glycoprotein, is more expressed in malignant glioma than in normal brain. It has been implicated in cell adhesion and migration in various cells, even if the function of this gene is not understood clearly. This gene has two isoforms of wild-type and splicing variant.

We revealed that only wild-type CD99 is expressed in human glioma cells and tissues. Through MTS, colony-forming, and anoikis assays, we found that wild-type CD99 in glioma cells has enough resistance for anoikis and ability to survive and grow in anchorage independence, as is opposed to previous reports in other tumors. Using tissue microarray, we validated that CD99 demonstrates higher expression in gliomas compared to non-neoplastic brain. We inhibited CD99 expression by siRNA and demonstrated decreased glioma cell migration and invasion. In contrast, when CD99 was over-expressed in glioma cells, we observed enhancement of cell migration and invasiveness. Orthotopic brain tumor model with nude mice harboring CD99 overexpressing glioma cells results in significant decreased survival rate, compared to control mice. Moreover, higher invasiveness is observed in overexpressing mice by pathological review.

Taken together, our findings suggest that CD99 play an important role in the migration and invasion of human gliomas. CD99 could be a target to inhibit migration and invasion especially in CD99-expressing gliomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2355. doi:10.1158/1538-7445.AM2011-2355
Abstract 2362: Antimetastatic potential of the antiprogestin mifepristone in cancer cells
Authors: Brandhagen BreeAnn N. , Tieszen Chelsea R. , Telleria Carlos M. , Goyeneche Alicia A.
Published: June 2012

For most cancers, it is metastasis and burden of the secondary tumors that account for nearly 90% of cancer-related mortality. The process of metastatic dissemination of tumor cells is extremely complex. Among the critical parameters involved in metastatic spread are cellular detachment from the initial tumor, migration, invasion and adhesion to secondary sites, all processes associated with changes in cell morphology caused by cytoskeletal rearrangements. Our laboratory previously demonstrated that the synthetic antiprogestin mifepristone (MF) inhibits the growth of cancer cells of reproductive and non-reproductive origin. We also observed that in response to MF, the cells displayed drastic changes in morphology. Due the impact of cell structure on the metastatic process, this study questioned whether MF has anti-metastatic potential deregulating key processes required for metastatic dissemination. Cancer cell lines of the ovary (SK-OV-3), breast (MDA-MB-231), prostate (LNCaP), and brain (U87MG) were chosen from a panel of cell lines in which we previously observed cytostatic activity of MF associated with changes in cellular morphology. Cell cultures were exposed to vehicle or MF for 72 h at which point we examined the morphology of the cells, the distribution of cytoskeletal proteins actin and tubulin, the adhesion capacity to components of the extracellular matrix, and the cell migratory and invasive properties. MF induced distinct morphology changes after 48 h exposure, independent of the density of the cell culture population. Interestingly, MF induced similar morphology changes in all cell lines, which included shrinkage of the cell body, long, thin cellular extensions, and a loss of cell-to-cell connections. Morphology changes were associated with rearrangement of actin and tubulin filaments. Upon removal of MF, cell morphology and proliferation returned to that of untreated cultures in all cell lines expect for LNCaP. For all cell lines, MF caused accelerated trypsin-induced detachment and decrease in cell adhesion to various extracellular matrix substrates suggesting that MF may impair re-adhesion of cells en route to colonize a secondary location. Moreover, MF significantly inhibited the ability of cells to migrate in wound healing and Boyden chamber assays, as well as their capacity to invade through extracellular matrix-coated polycarbonate membranes. Overall this study suggests that antiprogestin MF could be of potential therapeutic use, not only as a cytostatic agent impairing tumor growth, but also to halt metastatic spread of tumor cells. (Supported by NCI grants K22CA121991 and K22CA121991S1).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2362. doi:10.1158/1538-7445.AM2011-2362
Abstract 2377: Variation in the BRCT1 domain of the DNA repair gene XRCC1 delays invasive breast cancer in mice
Authors: Pettan-Brewer Christina , Morton John , Coil Rebecca , Goh Jorming , Ladiges Warren C.
Published: June 2012

The breast cancer susceptibility protein BRCA1 C-terminal (BRCT) domain coordinates protein interactions in DNA repair and cell cycle control, and comprises over 100 member proteins. One of these, XRCC1, is involved in base excision repair and single strand break repair. In order to investigate the role of the XRCC1-BRCT1 domain in breast cancer, we generated a mouse line with a variation of leucine (L) from arginine (R) at position 360, and crossed it with PyMT transgenic mice with a highly penetrant metastatic breast cancer phenotype. PyMT mice with advanced tumors expressing the L360R variant (N=20) had an average tumor volume of 360±57 mm3 compared to an average tumor volume of 730±69 mm3 for PyMT/wild type littermates (N=20), significant at p less than 0.01. In addition, the L360R variant was associated with a significant decrease in metastatic foci in the lungs of PyMT mice, an increase in tumor cell apoptosis as measured by caspase 3 immunohistochemistry, a decrease in tumor cell proliferation as measured by ki-67 immunohistochemistry, and a decrease in PARP binding as determined by NADH depletion. These preliminary observations suggest that the XRCC1/BRCT1 domain is involved in invasive breast cancer, and that genetic variation of this domain can suppress the invasive phenotype by decreasing tumor cell proliferation and increasing apoptosis of tumor cells possibly in association with altered PARP binding and activity. The XRCC1 L360R mouse is a promising animal model to further investigate the potential of targeting the XRCC1/BRCT1 domain for adjunctive treatment of invasive breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2377. doi:10.1158/1538-7445.AM2011-2377
Abstract 2396: Function of HLTF in the development of colon cancer and in the maintenance of genomic stability
Authors: Sandhu Sumit , Wu Xiaoli , Mai Sabine , Ding Hao
Published: June 2012

HLTF (Helicase like Transcription Factor) is a DNA helicase protein which is homologous to SNF/SWI family. It has also been shown to be a functional homolog of yeast rad5, required for the maintenance of genomic stability through the ubiquitination of proliferating cell nuclear antigen (PCNA). Although the physiologic role of HLTF is largely unknown, inactivation of HLTF by promoter methylation has been demonstrated in more than 40% human colon cancers. In this study, we have applied mouse transgenic approaches to determine whether loss of HLTF function could be important for colorectal carcinogenesis.

HLTF knockout mice were created by the deletion of first 5 exons of HLTF genome. The complete loss of HLTF expression in these mice was confirmed by Northern blot and RT-PCR assays. HLTF knockout mice did not show any developmental defects within a 2-year observation, indicating that HLTF is dispensable for normal development, and loss of HLTF function alone is not sufficient to drive colon carcinogenesis. To determine whether loss of HLTF function could cooperate with other tumor suppressors in the formation of colon cancers, we have bred HLTF knockout mice with the mutant mice for p53 and Apc (adenomtous polyposis coli). In HLTF−/−/Apc+/− mice, the significantly increased formation of intestinal adenocarcinoma and colon cancers were observed. Although very few HLTF−/−/p53−/− mice developed colon cancers, these mice had increased incidence of lymphoma formation. This finding suggests that loss of HLTF function could not only promote colon cancer formation, but also for other types of tumor. Cytogenetic analysis of colon cancer cells derived from HLTF−/−Apc+/− mice demonstrated a high incidence of gross chromosomal instabilities, including Robertsonian fusions, fragments and aneuploidy. All these genetic alterations were not observed in the intestinal tumor cells from Apc+/−, implicating that loss of HLTF function could induce genomic instability which contributes to colorectal carcinogenesis. To further establish the role of HLTF in the maintenance of genomic stability, we have also derived HLTF deficient mouse embryonic stem (ES) cells, and demonstrated that HLTF−/− ES cells exhibited similar, but a milder genetic alterations as observed in HLTF deficient colon cancer cells. Taken together, our work provides direct genetic evidence that loss of HLTF function as observed in a majority of human colon cancers is able to promote colorectal carcinogenesis. This function could be mediated by the induced genomic instability.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2396. doi:10.1158/1538-7445.AM2011-2396
Abstract 2401: Loss of Nkx3.1 cooperates with Myc overexpression to promote prostate tumorigenesis
Authors: McKissic Sydika , Roh Meejeon , Abdulkadir Sarki
Published: June 2012

Nkx3.1 is a homeodomain-containing transcription factor essential for prostate development and differentiation while loss mediates prostate tumor initiation. In mice, conditional loss of Nkx3.1 results in prostatic epithelial hyperplasia and prostatic intraepithelial neoplasia (PIN) formation. This increases the susceptibility of mice with compound mutations to the development of invasive adenocarcinoma. Human clinical data suggests that loss of Nkx3.1 and Myc overexpression are strongly correlated with increased tumor stage. Furthermore, Myc overexpressing transgenic mice lose Nkx3.1 in the PIN to carcinoma transition suggesting that Nkx3.1 loss is distinct and may provide a selective growth advantage. To further evaluate the cooperation between Nkx3.1 and Myc, we generated transgenic mice with prostate specific loss of Nkx3.1 and Myc overexpression. These mice contain a latent, Cre-activatable Myc allele and Nkx3.1 floxed alleles. Prostate specific Cre results in concurrent deletion of Nkx3.1 and activation of Myc. We have found that the young trigenic mice show advanced lesions compared to controls. High grade prostate intraepithelial neoplasia (HGPIN) was evident as early as 15 weeks of age. By 35 weeks of age, these mice show multi-focal HGPIN lesions with micro-invasive cancer. An increase in proliferation and a modest increase in apoptosis are also seen. Complementary prostate regeneration studies further support this progressive phenotype. We hypothesize that Nkx3.1 and Myc are cooperating to promote prostate tumorigenesis by regulating important cancer genes. Future studies are being conducted to identify the mechanism of cooperation between Nkx3.1 and Myc.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2401. doi:10.1158/1538-7445.AM2011-2401
Abstract 2403: Characterization of p53 in transgenic mouse prostate carcinogenesis models
Authors: Spees Colleen K. , Thomas-Ahner Jennifer M. , Tan Hseuh-Li , Yu Jun-Ge , Smolinski Justin B. , Kovalenko Pavlo L. , Fleet James C. , Clinton Steven K.
Published: June 2012

Introduction: Well characterized rodent models of prostate carcinogenesis are critical for the preclinical evaluation of dietary and chemopreventative interventions. p53 signaling is a target for genetic dysregulation in transgenic murine models of prostate cancer. We have examined p53 expression by immunohistochemistry, temporally and spatially in the prostate, as a potential biomarker of efficacy for chemopreventive interventions.

Methods: p53 immunohistochemical staining in wild type mice and two murine models (TRAMP and Rbf/TgAPT121) of prostate carcinogenesis was evaluated. Prostates from male TRAMP (up to 28 wks) and Rbf/TgAPT121 mice (up to 32 wks) were histopathologically evaluated. Normal epithelium, prostatic intraepithelial neoplasia (PIN) and adenocarcinoma was evaluated by p53 (CM5) staining, proliferation index (Ki67) and apoptotic index (ApopTag). Digital image analysis was employed to assist in quantification of biomarkers.

Results: p53 nuclear staining is rare in wild type mouse prostate epithelium. In TRAMP, p53 accumulation is first seen in the nuclei of young mice in areas of hyperprasia and low grade PIN (11%), with greater staining as carcinogenesis progresses through invasive cancers (59%). In the Rbf/TgAPT121 model, a less aggressive model, the PIN can be further subclassified and the percentage of p53 stained epithelial cells in the PIN lesions increases with advanced grade (PIN I: 3%; PIN II: 9%; PIN III: 17%; PIN IV: 30%). p53 staining also correlates with the apoptotic index in the PIN lesions (PIN I: 7%; PIN II: 9%; PIN III: 12%; PIN IV: 13%). Androgen dependency of p53 staining was demonstrated in TRAMP mice, as castration attenuates and repletion of testosterone restores the p53 nuclear staining. p53 staining is predominately located in the basal layer of the glandular epithelium and similar indices were observed in the dorsal and lateral lobes of the prostate.

Conclusions: p53 nuclear staining in TRAMP and Rbf/TgAPT121 models of prostate carcinogenesis increases as a function of lesion grade and stage. The early expression of p53 in murine prostate carcinogenesis is in parallel with other biomarkers of progression, such as proliferation and apoptotic index. p53 staining may serve as an early indicator of effective chemopreventive agent intervention.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2403. doi:10.1158/1538-7445.AM2011-2403
Abstract 2407: Decreased inflammation and tumor stage in COX-2 knockout mice in the BBN model of bladder cancer
Authors: Choudhary Shilpa , Hegde Poornima , Pilbeam Carol , Taylor John A.
Published: June 2012

Introduction and Objective: Prostaglandins (PG) play a critical role in the development of solid tumors. Inhibition of cyclooxygenase-2 (COX-2), the committed step in PG synthesis, has been shown to impair tumor development in the BBN mouse model of bladder cancer. Because inhibitors of COX-2 have been reported to have anti-tumorigenic properties independent of COX-2, we evaluated the impact of gene disruption of COX-2 in the BBN mouse model.

Methods: We used 2 mo old male COX-2 knockout (KO) and wild type (WT) mice in an outbred CD-1 background. Mice were treated with (n=13-14 / group) and without (n=6/group) BBN in drinking water and euthanized after 25-27 wks. Bladders were harvested, weighed, bisected and step sectioned after formalin fixation. H&E stained sections were evaluated for pathologic stage/grade, percent involvement by invasive disease and degree of inflammatory infiltrate(grade 1-3).

Results: All animals receiving BBN had inflammatory changes at the time of euthanasia, with the grade being higher in WT (grade 2-3) than KO mice (grade 1-2). Six WT mice and 3 KO mice receiving BBN died, all within the first 18 wks of study, without gross evidence of tumor. Compared to previous studies with C57Bl/6 mice, tumor development was slower and burden was lower at 25-27 wks. All 7 BBN-treated WT mice had high grade invasive cancer at euthanasia: 6 pT1 and 1 pT3. Out of 11 BBN-treated KO mice at euthanasia, 10 had high grade cancer and 2 had invasive disease: 1 pT0, 6 CIS, 2 pTa (non-invasive) and 2 pT1. Mean bladder weight at euthanasia was elevated only in BBN-treated WT mice. There was a strong correlation between WT genotype and tumor stage by Pearson's R (0.74, p=0.03). The percent of tumor representing invasive disease was 0.31 in COX-2 WT mice vs 0.01 in KO mice (t-test, p=0.002).

Conclusions: Genetic disruption of COX-2 resulted in lower tumor stage and percent invasiveness in the BBN mouse model of bladder cancer. However, most COX-2 KO mice developed high grade disease, indicating disruption of COX-2 was not sufficient to prevent tumorigenesis. Reduced inflammation in the COX-2 KO mice may account for the decreased invasiveness.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2407. doi:10.1158/1538-7445.AM2011-2407
Abstract 241: Transcription factor RBP-J-mediated signaling regulates basal cell carcinoma growth
Authors: Yu Mei , Zloty David , Bell Robert H. , Haegert Anne , Carr Nicholas , Shapiro Jerry , Cowan Bryce , Warshawski Larry , McElwee Kevin J.
Published: June 2012

Hair follicles (HF) and basal cell carcinomas (BCCs) can be regarded as ordered and disordered skin appendages respectively. They may utilize similar molecular mechanisms of growth. We examined the similarities and differences in gene expression between BCCs and HFs to define common and unique signaling pathways that distinguish an ordered skin appendage from a disordered skin growth. Nodular BCCs, non-follicular skin epithelium, and HF between the sebaceous gland and bulb region were microdissected and examined using microarrays. Selected genes were validated using quantitative PCR, immunohistochemistry and in vitro studies. Two differentially expressed gene sets were identified by significance analysis of microarray (SAM) in BCC and HF versus skin epithelium respectively. Subsequently, multiple signaling pathway analyses were conducted. The results indicated that Notch and Hedgehog signaling pathways were active in the growth of both HF and BCCs. However, Notch signaling, including tumor suppressor genes NOTCH1, NOTCH2, ligands JAG1, JAG2, signaling inhibitor NUMB, and downstream Notch pathway genes DTX1, DTX2, RBP-J, LFNG, HR, and HES7, all showed significant differential expression in BCCs compared to HF. The data suggests downstream gene expression in the Notch signaling pathway is suppressed in BCCs. We tested the effect of transcription factor RBP-J and found transcription factor RBP-J-mediated signaling suppresses BCC cell growth and induces cell apoptosis in vitro. Our data suggest that RBP-J serves as a tumor suppressor in BCC and BCCs deficient in RBP-J lose tumor repression activity. Modulation of the Notch pathway may be a focus for the development of BCC treatments.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 241. doi:10.1158/1538-7445.AM2011-241
Abstract 242: The gliogenesis initiation factor Nuclear Factor IB induces differentiation and inhibits growth of glioblastoma
Authors: Stringer Brett W. , Day Bryan W. , Barry Guy , Piper Michael , Jamieson Paul R. , Ensbey Kathleen S. , Charmsaz Sara , Boyd Andrew W. , Richards Linda J.
Published: June 2012

Introduction: The phylogenetically-conserved vertebrate transcription factor, nuclear factor IB (NFIB), is a key component in the differentiation of astrocytes during the process of gliogenesis in the developing mammalian central nervous system; a process that goes awry following various genetic and epigenetic alterations during the genesis of glioblastoma (GBM), the commonest and most aggressive form of primary human adult brain cancer. We found expression of NFIB to be reduced in GBM compared to normal human brain tissue so investigated what effect increased expression of NFIB had on GBM.

Experimental procedures: Expression of NFIB was investigated by qPCR and western blot in normal human brain tissue, primary GBM surgical specimens and GBM cells lines. The Rembrandt database was interrogated for patient survival data relative to NFIB expression. GBM cell lines were derived from patient tumor tissue, following informed consent, and cultured under serum-free conditions to help preserve the phenotype of the original tumor. GBM cell lines were transfected with HA-tagged Nfib. Glial differentiation marker expression was investigated by qPCR and western blot. Stem/progenitor cell growth was investigated by neurosphere assay and by PKH26 staining. Cell proliferation was measured by MTS assay and Ki67 staining. Cell cycle analysis was performed by PI staining and FACS analysis. Apoptosis was investigated by cleaved caspase staining. Differential gene expression induced by Nfib expression was determined by microarray analysis. In vivo tumorigenicity was investigated using subcutaneous and intracranial xenografts in NOD/SCID mice.

Results: We found NFIB expression in both GBM surgical specimens and GBM cell lines to be reduced relative to normal brain tissue. Reduced NFIB expression was associated with poorer patient survival. Increased expression of Nfib in transfected GBM cell lines induced expression of glial differentiation markers, inhibited cell proliferation, reduced stem/progenitor cell growth, altered cell cycle progression, and inhibited tumor growth in murine models of GBM.

Conclusion: We have identified NFIB as a novel tumor suppressor gene in human GBM.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 242. doi:10.1158/1538-7445.AM2011-242
Abstract 2426: Differential effects of liver metastatic and brain metastatic subset of murine breast carcinoma cells
Authors: Erin Nuray , Tanrιöver Gamze , Kale Şule , DuymuŞ Özlem
Published: June 2012

Carcinomas are formed by heterogeneous group of cells which differ in their aggressiveness as well as metastatic potential. Understanding the characteristics of cells that form distant metastasis is crucial to design effective treatments. We previously isolated liver metastatic cells (4TLM) formed by 4THMpc murine breast carcinoma using an orthotopic model. Here we observed that 4THMpc cells also generated macroscopic brain metastasis. Cells from brain metastasis were cultured in-vitro and named as 4THM-BM in order to determine differential characteristics of brain metastatic cells from liver metastatic cells. Specifically following features were compared: 1. Metastatic characteristics after mammary pad (MP) injection; 2. The effects of conditioned medium (CM) prepared from ex-plants of primary tumors as well as cancer-associated fibroblasts (CAF) on tumor cell growth. 3. MIP-2 levels found in CM of ex-plants of primary tumors and associated fibroblasts. Methods: 4TLM and 4THM-BM cells (100000 cells /mouse) were inoculated into the right upper MP of 8-10 weeks old female Balb-c mice. Necropsies were performed 25-27 days after injection. CM of ex-plants from primary tumors and surrounding fibroblasts were prepared. Results: Cells that were previously metastasized to liver (4TLM) produced significantly more macroscopic (52.2± 8.5 vs. 16.3 ±2.84 as mean number of nodules per animal) as well as microscopic lung metastasis (12.75 ±4.26 vs. 3.17±1,1 as mean % area involved with metastatic cells per animal) compared to brain metastatic cells (4THM-BM). Similarly number of macroscopic liver metastases was significantly higher (3.4± 0,6 vs. 7,3 ±1,23 as mean number of nodules per animal) in 4TLM injected animals. CM from ex-plants of 4TLM and 4THM-BM primary tumors as well as from corresponding CAF significantly enhanced growth of 4TLM cells but not 4THM-BM cells. MIP-2 levels in CM from ex-plants of CAF were higher from CM of 4TLM and 4THM-BM primary tumors. These results not only demonstrate the heterogeneous nature of the carcinomas but also suggest that cells that metastasize to liver are more aggressive in nature and produce more visceral metastasis. MIP-2, murine equivalent to human IL-8, is involved in angiogenesis as well as inflammation and reported to be highly expressed in metastatic cells. Here we found that both primary tumors formed by metastatic cell lines and associated fibroblasts produce MIP-2 which may contribute to the metastatic behavior.

Study was supported by The Scientific and Technological Research Council of Turkey (TÜBITAK Grant no:109S449).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2426. doi:10.1158/1538-7445.AM2011-2426
Abstract 2436: Endothelial integrin α3β1 stabilizes TF antigen/Galectin-3 mediated metastatic cell adhesion to the endothelium
Authors: Glinskii Olga V. , Li Feng , Rittenhouse-Olson Kate , Barchi Joseph , Glinsky Vladislav V.
Published: June 2012

Metastatic cell arrest in distant organ microvessels constitutes an essential rate-limiting step in hematogenous cancer metastasis. Previously, in a series of works we demonstrated the importance of interactions between cancer-associated Thomsen-Friedenreich antigen (TF-Ag) and endothelial galectin-3 (Gal-3) in facilitating the initial carbohydrate-mediated tumor cell adhesion to distant organ microvasculature. However, it was unclear which molecules stabilize TF-Ag/Gal-3 mediated adhesion and how these interactions effect subsequent events in a metastatic cascade. Here, the results of a pull down assay using TF-Ag coated gold nanoparticles followed by Western analysis identified endothelial integrin α3β1 being physically associated with TF-Ag/Gal-3 complexes. Blocking α3β1 did not affect the initial tumor cell adhesion to the endothelium, but reduced >10-fold shear force necessary to displace cancer cells stably attached to endothelial monolayers, indicating that α3β1 stabilizes tumor-endothelial cell adhesion initiated by TF-Ag/Gal-3 interactions. Subsequent Quadrupole Time-of-Flight mass spectrometry analysis revealed additional 7 proteins forming a macromolecular complexes with TF-Ag, Gal-3 and α3β1 such as filamin B, talin, IQ motif containing GTPase activating protein 1, vinculin, α actinin 4, zyxin, and T-plastin, all of which map onto focal adhesion pathway. Our data demonstrate an induction of complex signaling pathways downstream of this macromolecular complex facilitating endothelial cell retraction and metastatic cell transendothelial migration. Thus, endothelial α3β1 mobilization downstream of TF-Ag/Gal-3 interactions stabilizes tumor-endothelial cell adhesion, initiates focal adhesion formation, and induces signaling pathways in endothelial cells promoting cancer metastasis.

This work was supported in part by Award Number 1I01BX000609 from the Biomedical Laboratory Research & Development Service of the VA Office of Research and Development to VVG and AHA National SDG 0830287N to OVG.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2436. doi:10.1158/1538-7445.AM2011-2436
Abstract 244: Lung adenocarcinomas show genetic-morphologic correlations in three dimensional culture
Authors: Matsubara Daisuke , Ishikawa Shumpei , Fukayama Masashi , Niki Toshiro
Published: June 2012

EGFR mutation is associated with papillary morphology suggesting a genetic-histologic link. In this study we analyzed the relationship between gene expression, mutation, and morphology on Matrigel. Based on EGFR and MET abnormality combined with gene expression pattern, lung adenocarcinoma could be classified into two groups: Group I (Bronchial epithelial phenotype) with high expressions of CK7, MUC1, TTF-1, and E-cadherin, and Group II (non-Bronchial epithelial phenotype) with high vimentin expression (Matsubara et al, Am J Pathol, 2010). On Matrigel, cell lines formed either round (n=30), grape-like (n=4) or stellate-shaped (n=6) colonies. Most of the Group I cell lines formed round colonies. Most of the cell lines that formed grape-like or stellate-shaped colonies belonged to Group II. Virtually all cell lines with EGFR, HER2, and MET gene abnormality belonged to Group I and formed round colonies. In contrast, KRAS mutants were heterogeneous and formed either round, grape-like or stellate-shaped colonies. All KRAS-mutated Group I cell lines formed round colonies, while KRAS-mutated Group II cell lines frequently formed non-round colonies. Strikingly, non-round colony formation was correlated with loss of LKB1 expression in KRAS mutants. Thus, lung adenocarcinomas show genetic-morphologic correlations in three dimensional culture. The results reinforce the genetic-histologic correlations previously documented in primary tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 244. doi:10.1158/1538-7445.AM2011-244
Abstract 2440: Inhibition of SDF-1/CXCR4 dependent-metastases by the treatment with metabotropic glutamate receptor 5 (mGluR5) antagonists in oral cancer
Authors: Uchida Daisuke , Kuribayashi Nobuyuki , Tamatani Tetsuya , Nagai Hirokazu , Miyamoto Youji
Published: June 2012

Previously, we identified a metastasis-candidate gene, metabotropic glutamate receptor (mGluR)-5 by using an oral cancer cell line, B88-SDF-1, which have an autocrine stromal cell-derived factor (SDF)-1/CXCR4 system and distant metastatic potentials in vivo. In this study, we examined the effect of mGluR5 antagonists on the SDF-1/CXCR4 dependent-metastases in oral cancer. We used 2 kinds of mGluR5-specific antagonists, 2-Methyl-6-(phenylethynyl)pyridine (MPEP) and 3-((2-Methyl-1,3-thiazol-4-yl)ethynyl)pyridine (MTEP). These antagonists did not affect the growth of B88-SDF-1 cells, but did inhibit the enhanced-migration of the cells. Under these conditions, actin polymerization and pseudopodia formation were also inhibited by the treatment with MPEP and MTEP. In order to confirm the drug toxicities, these antagonists were intraperitoneally injected into wild-type mice for 4 weeks. All the mice injected with MPEP (5 mg/kg) and MTEP (5 mg/kg) did not show side effects, such as hematotoxicity, allergic reaction and body weight loss. When the B88-SDF-1 cells were transplanted into the vein of nude mice, these antagonists significantly inhibited the lung metastases of the cells. These results suggested that blockade of mGluR5, using antagonists such as MPEP and MTEP, may be effective at preventing metastasis in patients with CXCR4-related oral cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2440. doi:10.1158/1538-7445.AM2011-2440