bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract B3: Potential new mechanism whereby IGFBP-5 contributes to cancer progression: Binding of GADD34 and enhancement of eIF2α dephosphorylation
Authors: Ho Penny PY , Grkovic Steve , Firth Sue M. 
Published: May 2012

Abstract
Insulin-like Growth Factor Binding Protein (IGFBP-5) has a well-defined role in normal mammary gland physiology and has recently been shown to play an important role in the biology of breast cancer; however the IGF-independent mechanisms of action remain obscure. Over-expression of IGFBP-5, but not the addition of the exogenous protein, has been shown to inhibit the growth of breast cancer cells in vitro. This suggests novel intracellular mechanisms where IGFBP-5 interacts with unidentified binding partners and thereby modulates the proliferation and survival of breast cancer cells. To address this issue, we performed a yeast two-hybrid screen utilizing a normal human mammary gland cDNA library and full-length IGFBP-5 as the bait. The pre-eminent binding partner identified was Growth Arrest and DNA Damage-inducible protein 34 (GADD34), which assembles an eukaryotic initiation factor 2 alpha subunit (eIF2α) dephosphorylation complex at the endoplasmic reticulum (ER). We have confirmed that IGFBP-5 and GADD34 interact through coimmunoprecipitation experiments using ectopically expressed IGFBP-5 and GADD34 in mammalian cells. We have also shown that IGFBP-3, which belongs to the same protein family as IGFBP-5, interacts with GADD34 through coimmunoprecipitation studies. Furthermore, IGFBP-5 or IGFBP-3 expression was found to dramatically increase GADD34 protein levels as demonstrated through immunoblot analysis. However, IGFBP-2, another protein related to IGFBP-5, did not interact with GADD34 or increase its protein expression. Because GADD34-mediated eIF2α dephosphorylation is required to promote resumption of protein translation after periods of ER stress, we next examined eIF2α phosphorylation levels in the presence and absence of IGFBPs. Expression of IGFBP-5 or IGFBP-3 decreased the levels of phosphorylated eIF2α, suggesting that the interactions diminish the magnitude and/or duration of ER stress-induced inhibition of translation. Our results demonstrate not only a physical interaction between IGFBP-5 or IGFBP-3 and GADD34, but also show that these IGFBPs appear to increase the stability and activity of GADD34. This discovery suggests a novel role for IGFBP-5 and IGFBP-3 in the regulation of a protective mechanism induced by ER stress. Since ER stress is one of the hallmarks of solid tumor progression, by modifying the tumor response to hostile microenvironments, it is likely that these IGFBPs can have significant impact on cancer growth, survival, and apoptosis.
Citation Information: Cancer Res 2009;69(23 Suppl):B3.
Abstract B31: B-raf deletion and pharmacologic inhibition enhance K-ras-driven tumorigenesis
Authors: Murriel Christopher L. , Kenski Denise , Yuan Wenlin , Chen Honglin , Hatzivassiliou Georgia , Hoeflich Klaus , Wallin Jeffrey , Belvin Marcia , Liang Yuxin , Seshagiri Somasekar , Davis David P. , Johnson Leisa 
Published: May 2012

Abstract
The Ras GTPase family controls numerous downstream signaling cascades in response to signals that regulate cellular processes including proliferation and survival. While Ras is one of the most prevalent targets for gain-of-function mutations in human tumors, questions remain regarding how the Ras effector pathway functions in mutant K-ras-driven tumorigenesis. Since an important function of K-ras involves B-Raf activation within the canonical MAPK signaling pathway, we initiated a study to determine B-raf's role in the context of mutant K-ras-driven tumor promotion and maintenance.
We began by delivering adenovirus expressing the Cre recombinase to the lungs of genetically engineered mice possessing a conditional K-rasG12D allele (K-rasLSL-G12D) and either 0, 1 or both copies of the B-raf gene flanked by LoxP sites (B-rafCKO). This procedure results in expression of mutant K-rasG12D in the presence or absence of one or both B-raf alleles deleted within the mouse lung. Surprisingly, we observe that B-raf deletion significantly enhances lung tumor number and burden and decreases overall survival. When we used a highly specific small-molecule inhibitor that targets B-raf in a murine non-small cell lung carcinoma line harboring the K-rasG12D mutation, we observed an increase in cell proliferation and soft agar colony formation. Further investigation revealed that treating K-rasG12D expressing cells with the B-raf inhibitor enhanced MEK and Erk phosphorylation. Therefore, our data suggests that while B-raf deletion does not inhibit K-ras-driven tumor initiation and disease progression, its presence may play a pivotal role in establishing negative feedback regulation of constitutive mutant K-ras activity.
Citation Information: Cancer Res 2009;69(23 Suppl):B31.
Abstract B35: Dynamics of MHC class II tumor antigen presentation is critical for effector antitumor CD4+ T-cell generation and tumor rejection
Authors: Bonorino Cristina , Maito Fabio , Souza Ana Paula 
Published: May 2012

Abstract
Absence of CD4+ T cell help leads to defects in CD8+ T cell memory responses.Although this phenomenon is well characterized in infectious immunity, little is known about the generation of anti-tumor CD4+ T cell responses in vivo, or even how relevant is T cell help for tumor elimination. CD4+ T cells appear differ from CD8+ T cells in their requirements for differentiation, the dynamics of encounter with antigen being critical in this process. We analyzed the relevance of MHC class II antigen presentation for tumor rejection, developing a B16EaRFP line to study class II restricted tumor antigen presentation and the cognate CD4+ T cell help in vivo. We found that, in a tumor heavily infiltrated by dendritic cells (DC), a progressive decline in different cell subpopulations is observed in draining lymph nodes (DLNs), suggesting an interruption of local cell traffic. Artificial increase in precursor frequency of tumor specific CD8+ T cells by adoptive transfer of naïve OT-I cells in C57Bl/6 mice injected with B16OVA tumor was able to delay tumor growth, but was not sufficient to block it completely. While tumor antigen presentation in class II molecules was detected in the DLN, and could be increased by delivery of necrotic or apoptotic tumor cells, tumor specific CD4+ T cells showed proliferation arrest in the DLN. Tumor growth was blocked only when dendritic cells (DCs) were activated by intra-tumoral LPS injection, reestablishing cell traffic to the LN, resulting in expansion of INF-γ producing anti-tumor CD4+ T cells. Our results suggest that CD4+ T cells are a major target of tumor mediated antigen presentation interruption, and that CD4+ T cell help is necessary for rejection of an established tumor. These observations have major implications for the design of cancer immunotherapy strategies.
Citation Information: Cancer Res 2009;69(23 Suppl):B35.
Abstract B38: Silencing of mortalin, the mitochondrial hsp70, as a novel anticancer treatment strategy
Authors: Masarwa Muhammad , Donin Natalie , Ziporen Lea , Fishelson Zvi 
Published: May 2012

Abstract
A handful of monoclonal antibodies have recently been introduced in treatment of hematopoietic malignant neoplasms and solid tumors. The complement system, an important arm of the immune system, is activated following anti-tumor antibody treatment. Complement proteins assemble to form channel-like pores called MAC (Membrane Attack Complexes) that inflict damage to target cells. Unfortunately, cancer cells have many defensive strategies against antibody-dependent complement-mediated cytotoxicity, including MAC shedding by exocytosis. Our recent results have implicated mortalin, the mitochondrial heat shock protein 70, in the removal of the MAC from the cell surface. Mortalin is known to have tumor promoting and anti-apoptotic effects. Our research hypothesis states that mortalin is a vital protector of tumor cells from immune damage. In order to examine this hypothesis, we set up two mouse models of cancer, of K562 erythroleukemia cells in nude mice and EL4 T-lymphoma cells in C57BL/6 mice. Mortalin siRNA was administered to tumor bearing mice with or without concomitant tumor directed antibodies. First, in vitro experiments showed that mortalin siRNA acts synergistically with anti-cancer antibodies in lysis of tumor cells. In vitro mortalin silencing in tumor cells by siRNA and then injection of the cells into mice demonstrated reduced tumor growth, relative to cells pretreated with scrambled siRNA. Mortalin silencing in vivo by intratumoral or intravenous injection of mortalin siRNA led to a significant decrease in tumor growth rate. Furthermore, mortalin siRNA also reduced tumor blood supply and caused tumor necrosis. Growth rate of EL4 and K562 tumors was also significantly inhibited by injection of the mortalin inhibitor MKT-077. Mortalin inhibition in tumors may thus be regarded as a novel adjuvant treatment to immunotherapy.
Citation Information: Cancer Res 2009;69(23 Suppl):B38.
Abstract B4: Specificity of ▵Np63α-repressed target genes is determined by changes in p63-interacting proteins
Authors: Ramsey Matthew R. , Ory Benjamin , He Lei , Ellisen Leif W. 
Published: May 2012

Abstract
The transcription factor p63 has been reported to have many functions, including regulation of proliferation, suppression of apoptosis, suppressing differentiation, and maintaining stem cell self-renewal. These various functions are carried out by activation or repression of a diverse set of target genes. However, the mechanisms that lead to selective regulation of target genes have remained elusive. In human Squamous Cell Carcinoma (SCC) ▵Np63α is expressed at high levels, and serves to block TAp73-mediated induction of pro-apoptotic target genes such as Puma. Our previous work has shown that ▵Np63α and TAp73 physically associate, and that p63 binds to the promoter of TAp73-activated target genes such as Puma. We have constructed a panel of p63 point and deletion mutants in order to examine the role of different functional domains in p63-mediated repression. Using Luciferase reporter assays and a physiologic reconstitution model with mutant p63 in SCC cells, we have identified mutants defective in suppression of apoptosis and TAp73 target genes. Examination of point mutants defective in DNA binding, but able to bind to TAp73 normally has revealed a requirement for DNA binding in suppression of TAp73 target gene activation and apoptosis, suggesting that ▵Np63α-TAp73 heteromeric complexes on DNA are repressive. We have also performed a si-RNA based screen for regulators of p73 activity, and identified Daxx as a p63-interacting protein involved in the repression of p73 activity in SCC cells. These data demonstrate that ▵Np63α functions as part of a repressive complex to regulate pro-apoptotic target genes in human SCC. Thus, ▵Np63α plays a crucial role in maintaining the survival of SCC cells and promoting tumorigenesis.
Citation Information: Cancer Res 2009;69(23 Suppl):B4.
Abstract B45: Epigenetic signature of human breast cancer subtypes
Authors: Badola Sunita , Parker Alexander , Campbel Mara , Boedigheimer Michael , Grubinska Barbara , Turci Susan , Tsui Kim , Damore Michael A. , Wilson Cindy A. 
Published: May 2012

Abstract
Gene expression profiling has been used to extend classification of human breast cancers into five main sub-types having distinct clinical outcomes. The established prognostic markers ER, PR and HER-2 remain prominent components of this refined scheme that delineates luminal (luminal A, luminal B and HER-2) from non-luminal (basal/BRCA1 mutant and normal-like) breast cancers. Gene expression combined with immunohistochemical localization of various tumor markers in the normal mammary gland support the hypothesis that luminal and non-luminal breast tumors originate from different cell types. In the current study we explore epigenetic determinants of the different breast cancer sub-types using the genome-wide DNA methylation patterns (Illumina Infinium) of fifty human breast cancer cell lines. Principal component analysis of baseline gene expression profiles (Agilent) showed a clear separation between the luminal and non-luminal sub-types in agreement with previously published studies using a largely overlapping panel of breast cell lines. Comparative marker selection analysis using the GenePattern analysis package was then performed independently on the DNA methylation and gene expression datasets. When the results of these two analyses were compared, a statistically significant reciprocal overlap of the selected markers was observed. These results strongly suggest that a large fraction of the most robust luminal gene expression classifiers are silenced by promoter methylation in non-luminal cells, and conversely that many of the best non-luminal classifiers are silenced in luminal cells. The classifier genes indentified in the breast cancer cell line panel include the most highly referenced luminal (KRT8, ESR1, AR, TFF1, TFF3, TOB1 and XBP-1) and non-luminal markers (DSC3, VIM, SFRP1, KRT5, KRT16, KRT17, and RUNX3 that have been used to classify primary breast cancers. These striking differences in DNA methylation patterns strengthen the hypotheses that luminal and non-luminal sub-types arise from different precursor cells and that their transcriptional programs may be largely established during the process of cell specification in the mammary epithelial bi-layer. Since CpG island methylation is stable and robustly preserved in archival tissues, it is likely that a methylation signature comprised of a few key genes could be used to, for example, identify poor-prognosis, basal-type breast cancers. It is also possible that such a test would be more quantitative, accurate and reproducible than the “triple-negative” diagnosis currently based on the absence of immumohistochemical staining for ER, PR and HER-2.
Citation Information: Cancer Res 2009;69(23 Suppl):B45.
Abstract B48: Germline mutations in BRCA1 and BRCA2 as predictive biomarkers of therapy response
Authors: Tassotto Mary Lynn , Sutherland Lindsay , Th'ng John 
Published: May 2012

Abstract
The results of in vitro chemosensitivity studies have shown some consistency with response to chemotherapy in patients, indicating that this approach may be used to identify patients that are resistant to conventional chemotherapy regimens. However, this approach is not routinely used in an adjuvant or neoadjuvant setting as a treatment strategy. Treatment selection for breast and ovarian cancer is currently based on pathological classification of the tumour, without assessment of in vitro chemosensitivity. BRCA1 and BRCA2 germline mutations are considered to be both prognostic cancer biomarkers and predictive biomarkers of breast and ovarian cancer susceptibility; however, their utilization as predictive biomarkers of therapy response has not been investigated. Expression levels of BRCA1 modulate chemosensitivity in breast cancer cell lines, and epigenetic regulation of BRCA1 leading to decreased BRCA1 expression is often observed in sporadic breast cancer. We have hypothesized that cells with mutations in BRCA1 or BRCA2 may be differentially sensitive to DNA damaging agents used in cancer therapy due to decreased levels of BRCA1 or BRCA2 gene expression in these cells. Using lymphocytes isolated from individuals with germline mutations in BRCA1 and BRCA2, we will present results showing the in vitro response to ionizing radiation and chemosensitivity to Paclitaxel and Doxorubicin, two chemotherapeutic agents commonly used in the adjuvant systemic treatment of breast cancer.
Citation Information: Cancer Res 2009;69(23 Suppl):B48.
Abstract B54: Identification of the PI3K-mTOR pathway as regulator of mutant p53 turnover in human cancer cells
Authors: Bug Monika , Dobbelstein Matthias 
Published: May 2012

Abstract
The tumor suppressor p53 plays a crucial role in our cells to induce cell cycle arrest or apoptosis in response to different kinds of genotoxic stresses. In 50% of all solid tumors, p53 is mutated by a single point mutation within the DNA-binding domain and thereby loses its activity as a tumor suppressor. However, at the same time, mutant p53 actively contributes to chemoresistance and tumor progression through a gain of function (Dittmer et al., 1993). It was previously shown that mutant p53 highly accumulates in tumors of mice with targeted germ line mutations of the p53 gene (knock-in), whereas normal cells of these mice express it at low levels (Lang et al., 2004; Terzian et al., 2008). This argues that the deregulation of tumor-specific signaling cascades leads to an accumulation of mutant p53 and subsequently to tumor progression.
Our study aims at identifying these signaling pathways that contribute to the regulation of mutant p53 expression, and to describe their mechanisms. We performed quantitative analysis of mutant p53 protein levels in a cancer cell line after the individual knock-down of all known human kinases using automated microscopy (high content cell screening).
The bladder carcinoma derived cell line 5637 (carrying the p53 R280T mutation) was transfected with three different synthetic siRNAs targeting each known kinase or kinase regulator in a 96-well format, and p53 was detected by automated immunofluorescence microscopy. In each of the resulting 2400 cell populations the p53 expression levels were determined for at least 500 individual cells. Alterations in the distribution of p53 staining intensities were assessed for each kinase relative to non-targeting control siRNAs. The results of the kinases, that lead to the strongest up- or down-regulations of mutant p53 in the screen, were confirmed in different cell lines (U251 and U373, both derived from glioma, with the p53 hot spot mutation R273H), by immunofluorescence and immunoblot analysis.
We identified a group of kinases currently known to act in the PI3-kinase → mTOR pathway as potential regulators of mutant p53. The knock-down of these kinases resulted in a decrease of mutant p53 expression. Studying the mechanism of mutant p53 regulation, we performed quantitative real-time PCR after siRNA transfection and found no significant impairment of the p53 transcription. On the contrary, posttranslational modifications (phosphorylation and acetylation) of mutant p53 were decreased after knock-down of the identified kinases, even more strongly than the overall levels of the protein. These observations support the idea that the degradation of mutant p53 is decreased in cancer cells through alterations of PI3K-mTOR pathway associated kinases, resulting in p53 accumulation and oncogenic gain of function.
Citation Information: Cancer Res 2009;69(23 Suppl):B54.
Abstract B56: Identification of genes involved in mammary tumor susceptibility and resistance in two strains of mice
Authors: Griner Nicholas B. , Compton Shannon L. , Dickinson Ellen S. , Boehringer Michael , Wiesmüller Lisa , Jerry D. Joseph 
Published: May 2012

Abstract
Loss of p53 can lead to a variety of cancers, including breast cancer. Mice heterozygous for the p53 gene (designated Trp53+/−) develop spontaneous mammary tumors, but this depends on the strain background and has been linked to a locus on chromosome 7 (designated SuprMam1). Mammary tumors are common in BALB/c-Trp53+/−females, but are rare in C57BL/6-Trp53+/− mice. Prevalence of genomic instability appears to contribute to the phenotype as loss of heterozygosity (LOH) through mitotic recombination is significantly more common among tumors arising in BALB/c-Trp53+/− mice compared to C57BL/6J-Trp53+/− mice. This increased LOH in BALB/c-Trp53+/− tumors was shown to be due to recombination events. The Balb/c strain has been shown to have a deficiency in non-homologous end joining (NHEJ) of DNA double strand breaks (dsb), however, this does not account for the increase of LOH events in tumors. Our hypothesis is that BALB/c-Trp53+/− mice are more susceptible to mammary tumors due to impaired dsb repair leading to LOH. Using the COMET assay, we demonstrate that dsbs persist longer in BALB/c-Trp53+/− mouse embryonic fibroblasts (MEFs) compared to C57BL/6J-Trp53+/− MEFs. Similarly, co-localization of H2AX and the homologous recombination protein RAD51 remain at dsb longer in BALB/c-Trp53+/− MEFs compared to C57BL/6-Trp53+/− MEFs suggesting a decreased efficiency of homologous recombination. Using a GFP reporter assay, homology-directed repair was decreased by 36% in MEFs from BALB/c-Trp53+/− compared to C57BL/6-Trp53+/−. Palb2 is a candidate gene as it lies within the SuprMam1 interval and has been shown to contribute to heritable breast cancer. Although no coding SNPs were found within Palb2, preliminary results show the BALB/c-Trp53+/− cells have a 30% decrease in Palb2 RNA compared to C57BL/6-Trp53+/− cells as assayed by RT-qPCR. These results identify defects in homology-directed repair of dsb in BALB/c which are linked to susceptibility to mammary tumors in this strain. Experiments are in progress analyzing these features in congenic strains. The results will elucidate potential low penetrance modifiers that determine resistance to mammary tumors in both mice and humans which can be used as targets for therapy as well as breast cancer screening.
Citation Information: Cancer Res 2009;69(23 Suppl):B56.