bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 2240: Pre-clinical potency of INK128, a highly potent TORC1/2 kinase inhibitor in the HER2 amplified breast cancer model
Authors: De Pradip KR. , Sun Yuliang , Wu Hui , Rommel Christian , Yi Liu , Dey Nandini , Leyland-Jones Brian 
Published: August 2012

Abstract
Background: The PI3K-AKT-mTOR pathway is commonly activated in breast cancers due to the amplification of receptor tyrosine kinase (HER2), the mutation of PIK3CA or the loss of expression/function of PTEN. About 20% of HER2+ breast cancer patients have PIK3CA mutation, which has resulted in a trend toward shorter progression-free survival with trastuzumab-based therapy. Resistance (intrinsic or acquired) to trastuzumab is a significant clinical challenge, partly due to the overriding activation of PI3K-AKT-mTOR pathway. The PI3K pathway activation leads to stimulation of the key growth and proliferation regulatory kinase, mTOR which can be blocked by allosteric inhibitors (rapalogous) or by ATP-comptetive mTOR kinase inhibitor (TORC1/2). Purpose: We investigated the effects of INK128; a potent and selective TORC1/2 kinase inhibitor alone or in conjunction with trastuzumab in HER2 amplified breast tumor cell lines. We also sought to identify the molecular response determinants of mTOR kinase inhibitor predicting potent activity in HER2+/trastuzumab-resistant cell lines. Experimental Design: Here, we used the HER2 amplified trastuzumab-sensitive breast tumor cell line (BT474), the trastuzumab-resistant breast tumor cell line (BT474HR) and HER2 amplified/PIK3CA mutated breast tumor cell lines (HCC1954, UACC893). Results: We demonstrate that 1) HER2+ cells are sensitive to INK128 (IC50 ∼40nM), 2) INK128 is also effective in trastuzumab-resistant and PIK3CA mutaed/HER2+ breast tumor cells (IC50 ∼20 and 20-30nM respectively), 3) INK128 is mechanistically distinct from rapalogous by a more potent and comprehensive inhibition of PI3K-AKT-mTOR signaling network: a) avoids S6K inhibition-mediated feedback activation of PI3K-AKT and b) abrogates 4EBP1 phosphorylation; 4) ligand-induced HIF1α accumulation is inhibited by prior treatment of INK128, and 5) the combination of trastuzumab plus INK128 more effectively blocked PI3K-AKT-mTOR signaling and HIF1α accumulation both in parental and trastuzumab-resistant cells. Conclusion: Our results suggest that i) therapeutic targeting of PI3K-AKT-mTOR signaling should be effective in abrogating resistance to trastuzumab therapy in HER2+ breast cancer, ii) combination of trastuzumab and INK128 is a promising treatment approach for HER2 overexpressed as well as trastuzumab-resistant breast cancer patients and iii) dual TORC1/2 inhibitor also mitigates the feedback activation of AKT caused by selective inhibition of mTORC1, and because of this, INK128 may yield greater therapeutic benefit in breast cancer patients.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2240. doi:1538-7445.AM2012-2240
Abstract 2242: The MAPK and PI3K signaling pathways in breast cancer: Crosstalk mechanisms and the effect on cell proliferation
Authors: Monlish Darlene A. , Cavanaugh Jane E. 
Published: August 2012

Abstract
The purpose of the current study is to assess crosstalk between the mitogen-activated protein kinase (MAPK) and the phosphatidylinositol 3-kinase (PI3K) pathways in breast cancer. More specifically, the goal is to investigate the interaction between the extracellular signal regulated kinases 1/2 and 5 (ERK1/2 and ERK5) and Akt and how these kinases act in concert to promote sustained proliferation in a breast cancer model. The MAPK and PI3K pathways mediate numerous cellular processes, including differentiation, growth, proliferation, and survival. To date, most related studies have focused on assessment of the crosstalk mechanisms between ERK1/2 and Akt, while the interplay of ERK5 has remained largely unstudied. Breast cancer patients with increased levels of ERK5 have been found to have a poorer prognosis than patients with lower levels, warranting further investigation of this kinase. The interaction between ERK5 and ERK1/2, as well as between ERK5 and Akt, and how these signaling mechanisms act in conjunction to promote cell proliferation in breast cancer cells remains to be elucidated. The BT-474 (ER+/PR+/HER2+) and MDA-MB-231 (ER-/PR-/HER2-) human breast cancer cell lines were utilized to examine ERK1/2, ERK5 and Akt activation following treatment with ERK and Akt inhibitors or siRNA for these kinases, respectively. Western blot analysis was performed to determine expression and activation of the kinases. An MTT assay was used to assess changes in cell proliferation following kinase inhibition for 12, 24, 48, 72 and 96 hours. In vivo studies utilizing the transgenic MMTV-neu mouse model were also performed. In the BT-474 cell line, inhibition of Akt resulted in increased ERK1/2 and ERK5 expression and activation. Interestingly, the increases in both total and phosphorylated ERK1/2 and ERK5 following Akt-inhibition were dose-dependent, meriting further study. Excised mammary tumors from our animal model demonstrated an age-dependent increase in both ERK1/2 and ERK5. Notably, an age-dependent decrease in Akt was also observed, supporting the hypothesis that crosstalk between these pathways may exist. Future studies involving dual kinase inhibition are planned to further assess crosstalk and proliferation. The data obtained from these studies indicate the presence of crosstalk between the ERK1/2, ERK5 and Akt pathways. A better understanding of this crosstalk may lead to patient-specific drug regimens to decrease the cytotoxic effects of chemotherapeutics and improve patient mortality.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2242. doi:1538-7445.AM2012-2242
Abstract 2246: Combined grape polyphenols regulate mammalian target of rapamycin (mTOR) signaling via a dual mechanism of Akt inhibition and AMPK activation
Authors: Rivera-Rivera Amilcar , Castillo-Pichardo Linnette , Dharmawardhane Suranganie 
Published: August 2012

Abstract
Breast Cancer (BC) is the major prevalent cancer in women in the United States and Puerto Rico with metastatic breast cancer (MBC), often resulting in death. Recent studies have shown that the Akt/mammalian target of rapamycin (mTOR) pathway is a critical signaling cascade that drives MBC. We recently reported that a combined diet of grape polyphenols (resveratrol, quercetin and catechin: RQC) at equimolar concentration is effective at inhibition of mammary tumor growth and metastasis to lung and bones in nude mice. The objective of this study is to investigate a role for RQC in prevention of MBC via inhibition of the Akt/AMPK/mTOR pathway. We found that RQC at 5 μM each inhibits Akt and p70S6 kinase (downstream effector of mTOR) activities in a metastatic variant of the MDA-MB-435 cell line and MDA-MB-231 human BC cells. RQC at 5 μM demonstrated a 2-fold activation of adenosine monophophate-activated protein kinase (AMPK), an endogenous inhibitor of mTOR that acts opposite to Akt. To determine the contribution of individual resveratrol, quercetin, and catechin to the effect of combined RQC on mTOR signaling, MDA-MB-231 cells were treated with RQC at 1, 3 and 9 μM individually and in combination. Results show that at 1 μM, individual or combined RQC did not increase AMPK activity, while at 3 μM, only Quercetin increased AMPK activity. Individual compounds at 1 μM did not affect Akt activity, while combined RQC decreased Akt activity by 20%. At 3 μM, Akt activity is downregulated by all compounds individually and in combination with combined RQC exerting a higher inhibitory effect on Akt activity than individual compounds. In general, the combination of RQC was more efficient at inhibition of the Akt/mTOR pathway than individual compounds. Thus, RQC has the potential to be used alone or in combination with other cancer therapeutics to impede progressive BC. This study was sponsored by the Department of Defense/Breast Cancer Research Program award W81XWH-07-1-0330 to SD and the U54 grant No. CA-096297 to UPR Comprehensive Cancer Center.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2246. doi:1538-7445.AM2012-2246
Abstract 2249: Mutations to the GSK3β regulatory region of CTNNB1 increase TCF transcription activity and are resistant to degradation by GSK3β
Authors: Manring Heather R. , Antinozzi Peter A. 
Published: August 2012

Abstract
Hepatocellular carcinoma (HCC) is currently the third most common cause of cancer related mortality worldwide. As with many cancers, aberrant activation of Wnt/β-catenin signaling plays a critical pathogenic role in the disease. Somatic mutations in genes of the Wnt pathway are widely observed in tumor tissue, where it has been estimated that 15-20% of tumors possess activating mutations in beta-catenin (CTNNB1). The turnover of CTNNB1 protein is tightly regulated in cells by the degradation complex which includes the CK1α and GSK3β kinases in a complex with Axin and APC. Mutations within the CTNNB1 gene at the CK1α phosphorylation site (Ser45) and GSK3β phosphorylation sites (Ser33, Ser37, Thr41) are observed clinically and result in CTNNB1 protein accumulation and enhanced transcription of Wnt target genes. The goal of this study is to characterize additional clinically observed CTNNB1 mutations in HCC. It is hypothesized that mutations within the phosphorylation domain in exon 3 will impact the phosphorylation of these key regulatory sites of CTNNB1. To assess the impact of three observed CTNNB1 mutations (H36P, I35S and S23R) on Wnt signaling activity, mutant CTNNB1 constructs were first created and their impact on Wnt signaling was measured using a luciferase-based Wnt reporter assay in transfected HEK293T cells. Compared to the wild type CTNNB1 control, Wnt transcription activity in the presence of H36P, I35S, and S23R mutants were increased by 6.55 ± 2.08, 6.76 ± 2.44, and 2.36 ± 1.01 fold, respectively. This compared to a 5.91 ± 1.38 fold increase observed with the phosphorylation mutant S45P. To evaluate if these mutations had a measurable impact on GSK3β regulation of CTNNB1 levels, each mutant was coexpressed with a GSK3β expression vector and evaluated using the same Wnt reporter system. As anticipated, GSK3β coexpression reduced Wnt reporter activity of the CTNNB1 control by 3.6 fold ± .33 where the S45P mutant was not affected. Coexpression of GSK3β did not attenuate increased Wnt activity observed with the mutants H36P and I35S, however it did decrease activity observed with the S23R by a similar magnitude of the wild type CTNNB1 control (2.27 fold ± .93). These results describe the activating nature of H36P, S45P, and I35S which likely mediate their effect via impaired CTNNB1 phosphorylation. To a lesser extent, S23R activated Wnt activity, however, is subject to attenuation by GSK3β phosphorylation.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2249. doi:1538-7445.AM2012-2249
Abstract 225: Protein tyrosine kinase 6 directly phosphorylates p130 CRK-associated substrate to promote peripheral adhesion complex formation and cell migration
Authors: Zheng Yu , Tyner Angela L. 
Published: August 2012

Abstract
Protein tyrosine kinase 6 (PTK6) is a nonmyristoylated intracellular tyrosine kinase evolutionarily related to Src kinases. Aberrant PTK6 expression and intracellular localization have been detected in human prostate tumors. Although the majority of total PTK6 is localized within the cytoplasm of PC3 cells, a human prostate cancer cell line, we found that the activated pool of PTK6 phosphorylated on tyrosine residue 342 is localized at the membrane. Ectopic expression of membrane targeted PTK6 leads to enhanced activation of PTK6 at the membrane and the formation of 6 to 12 striking focal adhesion-like complexes localized at the periphery of each cell, termed peripheral adhesion complexes. Using confocal microscopy, we show that p130 CRK-Associated Substrate (p130CAS), FAK, Paxillin and Vinculin, as well as increased levels of tyrosine phosphorylated proteins and active PTK6 are localized within the induced peripheral adhesion complexes. We determined that peripheral adhesion complex formation is dependent upon PTK6 tyrosine kinase activity. We demonstrate that p130CAS is a novel direct substrate of PTK6 that works as a crucial adapter protein in PTK6 mediated formation of peripheral adhesion complexes. We found that activation of ERK5 signaling downstream of p130CAS is indispensible for this process. Knockdown of p130CAS attenuates oncogenic signaling induced by membrane targeted PTK6, including ERK5 and AKT activation. We show that formation of peripheral adhesion complexes is associated with enhanced cell migration. PC3 cells expressing membrane targeted PTK6 exhibit increased migratory ability, which can be impaired by knockdown of p130CAS or ERK5. In contrast, knockdown of endogenous PTK6 in PC3 cells reduces cell migration, which is accompanied with decreased phosphorylation of p130CAS and ERK5. Our study reveals a novel function for PTK6 at the plasma membrane, and suggests that a PTK6-p130CAS-ERK5 signaling cascade plays an important role in cancer cell migration and invasion.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 225. doi:1538-7445.AM2012-225
Abstract 2253: New targets of mTORC1 pathway in ER-positive cells
Authors: Sedletcaia Anna , Unger Helen A. , Plotzker Naamah , Holz Marina K. 
Published: August 2012

Abstract
mTOR complex 1 (mTORC1) regulates protein translation, autophagy, and cell growth. These cellular functions are often dysregulated in cancer, therefore identification of mTORC1 targets in control of these processes is of great interest for development of targeted cancer therapies. While rapamycin shown an effect in early clinical trials, the effectiveness of rapamycin in treatment of breast cancer has been variable. Estrogen receptor (ER) ≤ mediates the proliferative effects of estrogen and represents an important clinical target in treatment of ER-positive cancers. Tamoxifen is an anti-estrogen that has become the standard agent for the treatment of ER-positive breast cancer, where it acts as an antagonist. However, resistance to tamoxifen, and other endocrine or anti-estrogen therapies develops in many cases. One mechanism by which resistance develops is through phosphorylation of ERα, allowing it to act in estrogen-independent manner. Phosphorylation of Ser118 and Ser167 is important for transcriptional activity of ERα. The regulation of these sites is complex, involving inputs from the mTOR and MAPK signaling pathways. Importantly, ERα phosphorylation correlates with resistance to tamoxifen, and is a prognostic marker for disease progression and survival. Therefore, the goal of our study is to identify new targets of mTORC1 pathway important for growth and proliferation of ER-positive cells. Thus, inhibition of downstream components of the mTORC1 pathway together in combination with anti-estrogen therapy may prove to be an effective strategy in treatment of ER-positive cancer.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2253. doi:1538-7445.AM2012-2253
Abstract 2261: Heat-shock and autophagy in cancer: Novel role for DGK iota
Authors: Samarasinghe Buddhini N. , Wales Christina T. , Jacobs Aaron T. 
Published: August 2012

Abstract
Autophagy is a cellular process by which damaged proteins, protein aggregates, and organelles are sequestered by lysosome-like compartments and degraded. Autophagy contributes to quality control, cell death, and senescence. Defects in autophagy cause the accumulation of cellular damage, inhibit cell death processes, and promote an inflammatory state that facilitates tumor development. In contrast, the heat shock response, which is mediated primarily by the transcription factor, HSF1 is frequently enhanced in cancer. Despite this inverse relationship between autophagy and heat-shock in cancer, the direct influence of heat-shock and HSF1-mediated gene expression on autophagy has not been investigated. Our study suggests that heat-shock mediated gene expression suppresses autophagy in a human colorectal cancer cell line. Our data show that diacylglycerol kinase iota (DGK iota) expression is regulated by HSF1. DGK iota phosphorylates diacylglycerol, generating phosphatidic acid (PA), which is a potent suppressor of autophagy through its activation of the serine/threonine kinase mTOR (mammalian target of rapamycin). The aim of our study was to evaluate the influence of HSF-1-mediated gene expression on autophagy, and to examine the specific contribution of DGK iota in this effect. We utilized RKO colorectal cancer cells for this investigation and assayed autophagy using complimentary techniques, including: conversion of LC3-I to LC3-II, which was monitored by Western blot; and the formation of autophagic vesicles (autophagosomes), which were visualized by high-content screening confocal microscopy of cells harboring an LC3-GFP expression construct. The influence of basal and heat-shock (42°C)-simulated HSF1 activity on autophagy was analyzed using siRNA. Silencing of HSF1 led to a profound increase in autophagic markers, suggesting a role for HSF1-driven gene expression in the suppression of autophagy. In HSF1-deficient cells we also observed increased phosphorylation of p70S6K, which is a surrogate marker for mTOR activation. We confirmed that DGK iota mRNA and protein expression is mediated by HSF1. Using biochemical inhibitors of DGK activity, we observed a significant increase in autophagy, supporting our hypothesis that DGK iota participates in suppressing autophagy. Additional siRNA experiments of DGK iota are being performed to confirm this effect, along with measurements of cellular PA levels. Taken together, these results indicate an important role for HSF1 in the suppression of autophagy, and provides mechanistic insight into a potential, tumor-promoting role for DGK iota.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2261. doi:1538-7445.AM2012-2261
Abstract 2265: Embelin induces autophagy in ovarian cancer cells
Authors: Jehan Zeenath , Al-obaisi Khadija Abdul Rahman , Uddin Shahab , Al-Kuraya Khawla 
Published: August 2012

Abstract
Embelin is an active ingredient of traditional herbal medicine that exhibits cytotoxic effect and inhibits cell proliferation in various cancer cell types. It is a small molecular inhibitor of XIAP undergoing preclinical testing for its anticancer effects in many different cancer types. In this study we have investigated its role as a chemotherapeutic agent in a panel of ovarian cancer cell lines. Utilising the MTT assay and annexin staining we demonstrate that embelin exhibits a potent, dose dependent cytotoxic effect in human ovarian cancer cells. Surprisingly in contrast to the previously reported cytotoxicity of embelin in many other cancer cell types, treatment of ovarian cancer cells with embelin promoted autophagy and not apoptosis as evidenced by the death without caspase 3 activation and induces cellular swelling and cytoplasmic vacuolization with evident features of necrotic cell death. Treatment of ovarian cancer cells with embelin demonsrnstrated autophagic hallmarks, including an increased number of autophagosomes were observed through acridine orange staining. Embelin also induced cell autophagy, as demonstrated by the punctuate distribution of GFP-LC3, as well as the LC3 cleavage and beclin1 activation. Red green fluorescence in acridine orange stained cells by flowcyometry further confirmed the autophagy action by embelin treatment. Our findings demonstrate a new death pathway underlying the cytotoxic effect of embelin, but also suggest that targeting autophagy could augment the anticancer activity, providing the framework for further development of embelin as a new chemotherapeutic agent for ovarian cancer treatment.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2265. doi:1538-7445.AM2012-2265