bluuebunny/crossref_metadata_2025_split huggingface dataset:
P1-12-08: Induction of Adhesion Molecules on Target Cells Maximizes Antibody-Dependent Cell-Mediated Cytotoxicity of Anti-Her2 Therapeutic Antibody with High Fucose Glycan.
Authors: Jia X , Wong P-Y , Gazzano-Santoro H 
Published: June 2012

Abstract
Background: Antibody-dependent cell-mediated cytotoxicity (ADCC) is one pivotal effector function of certain therapeutic antibodies. ADCC is the lysis of antibody-coated target cells by effector cells expressing Fc receptors (FcR) that recognize the fragment crystallizable (Fc) region of the bound antibody. Higher fucosylated terminal glycan on Fc results in significantly lower ADCC activity because core fucose sterically hinders the binding of Fc region to FcR on effector cells. The role of adhesion molecules in ADCC is unclear but it has been reported that adhesion molecules help better orientation of effector cells (natural killer cells) toward adhesion molecule-coated beads. We hypothesize that a preferable association of effector cells and target cells mediated by adhesion molecules may compensate ADCC loss due to the steric hindrance of high core fucose on Fc. Recombinant humanized monoclonal anti-Her2 antibodies (anti-Her2 rhuMAb) at various fucosylation levels have been used in this study to evaluate the role of adhesion molecules in ADCC of antibodies with high fucose glycan.
Materials and Methods: Natural killer (NK) cell line transfected with FcR were used as effector cells. Her2-expressing breast cancer cell lines BT-474 and SK-BR3 were used as target cells. Adhesion molecules were induced in target cells by incubation with 50ng/ml of recombinant human tumor necrosis factor (TNFa) for 1–2 days. Anti-Her2 rhuMAbs with different Fc fucosylation (G0-F from &lt;1% to &gt;2%) were added in the co-culture of effector cells and target cells. ADCC activities were quantitated by target cells lysis assessment using Europium-TDA release assay.
Results: Untreated BT-474 or SK-BR3 target cells had very little adhesion molecule expression. A high fucosylated anti-Her2 rhuMAb induced 52% of target cell lysis, which represented two-thirds of cell lysis by a low fucosylated Ab. Increment of E:T ratio could not maximize ADCC activities. With TNFa treatment, target cell lysis was increased from 52% to 100% by high fucose rhuMAb. This enhancement of ADCC activities is probably due to the expression of adhesion molecules such as ICAM-1 and VCAM-1, which was shown to be up-regulated by 100-fold on target cells in the presence of TNFa. Our data suggested that ADCC activities in Her2-expressing breast cancer cell lines were HER2−specific, anti-HER2 rhuMAb-dependent and adhesion molecules were indispensible for the enhanced ADCC activities.
Discussion: rhuMAbs produced in Chinese hamster ovary (CHO) cell lines may contain various Fc fucosylation, hence various ADCC activities. Although up-regulation of adhesion molecules alone cannot lyse target cells directly, our data suggested maximized ADCC activities by rhuMAbs with high Fc fucosylation level could be achieved by induction of adhesion molecule expression on target cells.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-12-08.
P1-12-16: HER-2 Testing and Treatment – Is Age a Factor?
Authors: Singh JK , Ewing K , Howell S , Howe M , Cramer A , Bundred NJ 
Published: June 2012

Abstract
Aims: Elderly breast cancer patients have a poorer prognosis due to late diagnosis and sub-optimal treatment. In the UK, HER-2 testing was introduced in 2005 and we sought to determine whether HER-2 testing was performed on patients of all ages and whether HER-2 positive patients of all ages subsequently received Herceptin, from implementation in 2005 until 2008.
Methods: Review of all newly diagnosed breast cancers in women over 50 in our Unit between January 2005 and December 2008 was performed. Cases were identified from the hospital pathology electronic database. Patients were considered HER-2 positive if tissue sections scored 3+ on immunohistochemical analysis (intense membranous staining) or if gene amplification was detected using fluorescence in-situ hybridisation (FISH). Patients were eligible for treatment with Herceptin if the tumour exceeded 1cm in size or if they had lymph-node positive disease, per UK guidelines. Herceptin treatment was determined for all patients.
Results: In total 703 patients with a median age of 68 years (range 56 to 98) were identified and 628 (89.3%) underwent definitive surgery (mastectomies: 246; wide local excisions: 382) and axillary surgery. In total 371 patients (52.8%) underwent HER-2 testing by immunohistochemistry or FISH. HER-2 tests performed by year: 2005 (51/131; 38.9%); 2006 (59/187; 31.6%); 2007 (71/189; 37.6%) and 2008 (190/196; 96.9%). Fifty six patients (15.1%) were HER-2 positive by immunohistochemistry or FISH. Median age of HER2 positive patients was 66 years (range 57 to 98); median age of HER2 negative patients was 68 years (range 56 to 96). Forty four HER-2 positive patients (78.6%) were eligible for treatment with Herceptin; of these 28 patients (63.6%) were treated with Herceptin. HER2 testing, treatment with Herceptin and overall 5-year mortality rate for patients between 50 to 69 years and patients aged 70 years or older is summarised in Table1.

Twenty percent of patients aged between 50 to 69 years and 71.4% of patients aged 70 years or older eligible for Herceptin were not treated. Reasons for non-treatment included: multiple co-morbidities (4); treatment declined by oncologists (1); no referral to oncologists (1); death prior to treatment (2) and unknown (8). Median follow up time was 43.3 months (range 0.2 to 73.8). All cause mortality was higher in the over 70 year old breast cancer group (Log Rank Test p=0.001) regardless of HER-2 status.
Conclusions: National introduction of routine HER-2 testing for all newly diagnosed breast cancers led to a testing rate increase to almost 97%. Despite this only 63.6% of HER-2 positive patients eligible for treatment with Herceptin received it. Patients aged 70 years or older were less likely to receive Herceptin due to multiple factors.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-12-16.
P2-01-13: Impact of Erlotinib on MSC-Mediated TIC Expansion and EMT.
Authors: Lacerda L , Solley T , Debeb B , Xu W , Krisnamurthy S , Ueno N , Reuben J , Klopp A , Woodward W 
Published: June 2012

Abstract
Background: Recently, we have demonstrated that mesenchymal stem cells (MSC) and MSC secreted factors (MSC-CM) have a profound effect on tumor initiating cells (TIC) enriched mammosphere formation and latency of tumor xenografts formation from breast cancer cell lines. Furthermore these interactions increased the expression of epithelial mesenchymal transition (EMT)-associated proteins which are associated with tumor cell invasion and metastasis as well as the TIC phenotype. (Klopp, A. H. et al., 2010, PLoS One. 5, e12180). Our data suggest that the presence of MSC in the tumor microenvironment may increase metastases by conferring stem progenitor cell biology on more differentiated non-metastatic cells. In addition, preliminary data suggested MSC-CM upregulated EGFR signaling in breast cancer cells. Therefore, we hypothesized that inhibiting EGFR signaling with erlotinib (tyrosin kinase inhibitor) can suppress MSC-mediated TIC expansion and EMT.
Methods & Results: In order to demonstrate that erlotinib inhibits MSC-CM promoted expansion of TIC, we cultured breast cancer cells lines (SUM149, SUM159, SUM190, MDA-IBC3 and MCF-7) in anchorage independent conditions with MSC-CM and treated them with increasing concentrations of erlotinib. The efficiency of mammosphere formation was examined after 5 days. We found that erlotinib inhibited MSC mediated increase in mammosphere formation in triple negative cell lines SUM149 and SUM159, and HER2−positive cell lines SUM190 and MDA-IBC3, but not in ER-positive, erlotinib resistant MCF-7 cells. Furthermore, we evaluated the impact of erlotinib on cell cultures grown with breast cancer patient-derived fluids, such as seroma and malignant pleural effusions. We observed that the effect of erlotinib on mammospheres formation was attenuated by both types of patient fluids.
Discussion: Patients with triple negative breast cancer have the highest rates of metastases and no available targeted therapies for treatment. EGFR is expressed in a significant proportion of triple negative breast cancers, and recent clinical and preclinical studies suggest that EGFR may contribute to the metastasis or aggressiveness of triple negative breast cancer. Here we demonstrate that host and environmentally-derived factors are critical for determining resistance to erlotinib. In vivo studies regarding the ability of erlotininb to prevent MSC-enhanced TIC survival and metastases are underway.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-13.
P2-01-14: High Level of CD44 Expression and Soluble CD44 Secretion Mediate Trastuzumab Resistance in HER2 Positive Breast Cancer.
Authors: Jin Q , Baek JM , Boulbes DR , Esteva FJ 
Published: June 2012

Abstract
Background: Overexpression of HER2 occurs in 15–25% of invasive breast cancer. Trastuzumab, a humanized monoclonal antibody targeting the extracellular domain of HER2, markedly improves disease-free and overall survival in patients with HER2−positive breast cancer. However, resistance to trastuzumab develops in most patients with HER2−overexpressing metastatic breast cancer. Breast cancer stem cells have been shown to express HER2, although its role in the development or maintenance of stem cells is not well characterized. CD44 is one of the proposed markers for breast cancer stem cells. We investigated the role of CD44 in acquired trastuzumab resistance in HER2 positive breast cancer.
Methods: CD44 mRNA expression was measured by quantitative real-time PCR. The proportion of CD44 positive cells was measured by fluorescence-activated cell sorting (FACS). Soluble CD44 level was detected by enzyme-linked immunosorbent assay (ELISA). In vitro tumorigenicity was evaluated by soft agar colony formation assay. Cell invasion was tested by using a matrigel invasion assay. In vivo tumor growth was evaluated using a mammary fat pad xenograft mouse model.
Results: CD44 mRNA expression, CD44 positive cell population by FACS, and CD44 secretion were significantly increased in both SKBR3 derived trastuzumab resistant SKBR3 clone 3 and BT474 derived trastuzumab resistant HR20 cells compared to parental, trastuzumab sensitive cells. Consistent with the increase in CD44, trastuzumab resistant cells showed an increase in soft agar colony formation and cell invasion, which were blocked by siRNA-induced downregulation of CD44 expression. Tumor growth and incidence were enhanced in mice injected with trastuzumab resistant HR20 cells compared to the mice injected with trastuzumab sensitive parental BT474 cells. Knock-down of CD44 expression by the transfection with CD44-specific siRNA suppressed in vivo tumor growth in the mice injected with trastuzumab resistant HR20 cells. Finally, we found high serum CD44 levels correlated with advanced tumor progression in HER2 positive breast cancer.
Conclusion: Our findings indicate that high levels of CD44 mediate trastuzumab resistance in HER2 positive breast cancer cells as well as tumor progression in mice. Circulating CD44 is prognostic in patients with HER2 positive breast cancer. CD44 may be a therapeutic target for HER2 positive breast cancer.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-14.
P2-01-17: L1-CAM Promotes Adhesion of Breast Cancer Cells to the Endothelium.
Authors: Milde-Langosch K , Dippel V , Schröder C , Wicklein D , Hein S , Jänicke F , Müller V , Schumacher U 
Published: June 2012

Abstract
Background: Overexpression of the adhesion molecule L1 -CAM (L1) in breast cancer is correlated with nodal involvement, high grading, and a shorter recurrence-free and overall survival, but the mechanism leading to this effect is poorly understood. L1 is also expressed in endothelial cells promoting the interaction between lymphatic cells and endothelia via homophilic L1-L1 interaction. This mechanism might also mediate adhesion of breast cancer cells to endothelia and thus promote metastasis. To examine this likely role of L1, the impact of L1 expression in breast cancer cells on their adherence to human pulmonary microvascular endothelial cells (HPMEC) was investigated and potential L1 ligands on these cells were identified. Methods: MDA-MB231-Fra2 breast cancer cells expressing high levels of L1 were stably transfected with shRNA vectors to generate two clones with strong L1 downregulation (L1low clones). Adherence of these clones to endothelial cells was studied in dynamic cell-flow adhesion assays in channels seeded with HPMEC simulating in vivo-conditions. Rolling or adherent cells per minute were counted using CapImage software. For the cell-flow assay HPMEC were activated with TNFα. In order to identify the L1 ligand, endothelial cells were partly pre-incubated with blocking antibodies directed to the potential binding partners L1, ALCAM, ICAM-1 and E-selectin before interaction with tumor cells. In additional experiments, flow chambers were coated with recombinant L1-CAM, ICAM1, ALCAM or E-selectin, and the adherence of MDA-MB231 cells with high or low L1 expression to these proteins was investigated.
Results: Adhesion of MDA-MB231 cells to activated HPMEC was significantly higher in L1high cells compared to L1low clones, where the number of adherent cells was only 40–50% of the L1high control (p=0.025; p=0.035). Blocking experiments showed that the adherence of L1high cells could be reduced by antibodies directed to ALCAM (p=0.0007), but not to ICAM1. Anti-L1 antibodies had a significant effect only in passages of endothelial cells which showed L1 expression, whereas E-selectin blocking strongly diminished adherence of breast cancer cells irrespective of their L1 expression. In addition, L1low clones showed significantly lower adhesion to recombinant L1, ALCAM and E-selectin proteins compared to the parental cells.
Conclusion: Our experiments indicate that L1 expression in breast cancer cells leads to an increased adherence to activated endothelial cells via homophilic (L1-L1) or heterophilic (L1-ALCAM) interactions. This mechanism is a possible explanation for the increased metastatic potential and poor prognosis in L1-positive carcinomas observed in vivo. Our results suggest that this adhesion molecule might be a suitable target for therapeutic interventions.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-17.
P2-01-23: Seprase Promotes the Growth and Impairs the Migratory Ability of Breast Cancer Cells.
Authors: Jia J , Martin TA , Jiang WG 
Published: June 2012

Abstract
Background: Seprase or Fibroblast Activation Protein alpha (FAP-a) is an integral membrane serine peptidase. It has been shown that it plays an important role in tumour proliferation, migration, invasion and angiogenesis. However, previous work has not satisfactorily explained both the suppression and promotion effects that have been observed. Furthermore, the interaction of Seprase with other membrane molecules or pathway had not been well investigated. This study aimed to investigate the role of Seprase in human breast cancer. Methods: The full length of Seprase cDNA was isolated from normal human prostate tissue using RT-PCR and then cloned into an expression vector (pEF6-V5 His, TOPO). Following confirmation of successful cloning, the plasmid was transfected into the to human breast cancer cell lines, MCF-7 and MDA-MB-231. The function of Separase in these two cell lines was then analysed using cell function assays and ECIS, and the potential interaction with other signalling pathways explored.
Results: Compared with the MCF-7 wild type cells (MCF-7wt) and the MCF-7 cells transfected with pEF6/V5 empty vector (MCF-7pef), the MCF7 cells transfected with Seprase (MCF-7exp) had increased growth ability and impaired migratory ability. The growth rate of MCF-7exp cells was 1132.72% on day 5 while those of MCF-7wt and MCF-7pef control cells were 897.24% and 990.99% respectively (p=0.011 and 0.032). The resistance increase of MCF-7exp cells at 2 hours after wounding in ECIS assays was 478.22ohms compare with 1063.75 ohms and 1563.09 ohms in MCF-7wt and MCF-7pef control cells (p&lt;0.05). In MDA-MB-231 cells, over-expression of Seprase resulted in impaired motility. The resistance increase of the MDA-MB-231 wild type cells (MDA-MB-231wt), MDA-MB-231 cells tranfected with pEF6/V5 vector (MDA-MB-231pef), and the MDA-MB-231 cells transfected with Seprase (MDA-MB-231exp) at 3 hours after wounding in ECIS assay were 216.75ohms, 216.00ohms and 120.16ohms respectively. The growth of MDA-MB-231 cells and the adhesion and invasion ability of both MCF-7 cells and MDA-MB-231 cells were not dramatically influenced with Seprase expression. An inhibitor to focal adhesion kinase (FAK) restored the reduced motility ability of both MCF-7exp cells and MDA-MB-231 exp cells. However, inhibitors to PI3K, ERK and ROCK had no influence on it.
Conclusion: These results suggest that Seprase promotes proliferation but inhibits the migration of breast cancer cells by potentially regulating the FAK pathway.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-23.
P2-02-09: TP53 Mutation Patterns in Breast Cancer Subgroups.
Authors: Dumay A , Feugeas J-P , Wittmer E , Lehmann-Che J , Bertheau P , Espié M , de Cremoux P , André F , Sotiriou C , Pusztai L , de Thé H 
Published: June 2012

Abstract
Tumor protein 53 (TP53) is the most commonly altered gene in human cancers. In breast cancers, TP53 is mutated in approximately 30% of all cases, but this frequency fluctuates widely within the different molecular subclasses. Different types of mutations may be observed, such as substitutions (replacement of a nucleotide pair by another one), or complex mutations (deletions or insertions of one or more nucleotides). Mutation types may reflect mechanism of DNA lesion or DNA repair deficiencies. Furthermore, mutations can give rise to different effects such as truncating mutations leading to loss of function, or missense mutations often leading to dominant negative activity. Those mutation effects can be advantageous in tumorigenesis and thus can be subject to selective pressure.
Here we classified 572 breast tumors in three groups, according to microarray data: luminal, basal and molecular apocrine. TP53 status was assessed by a yeast-based functional assay (FASAY) and cDNA sequencing. We then assessed whether any feature of TP53 mutations would be preferentially associated to a specific subtype of breast cancer.
- In term of TP53 mutation frequencies, as expected, lowest frequency was observed in luminal subgroup (26%) and highest in basal (90%) and molecular apocrine (70%). Notably, much higher rate of TP53 mutations occurred in luminal B subgroup (41%) than in luminal A one (17%), suggesting that TP53 may be an important feature in progression from luminal A to B.
- In term of mutations types, luminal tumors showed high frequency of substitutions, while molecular apocrine and basal presented increased rate of deletions and insertions, reflecting probably increased rate of DNA breaks. This suggests that same mutational events may occur in basal and molecular apocrine tumors.
- In term of mutation effects, we found high frequency of missense mutations in luminal tumors (notably AT to GC) and much higher rate of truncating mutations in basal tumors. These observations point to an existence of different selection pressure in each of them, such as a strong pressure for P53 mutations with potential dominant negative inhibition of P73/P63 (recently shown to favor invasion), in luminal tumors. Collectively, these results point not only to different mechanisms of P53 gene inactivation, but also different functional consequences among the different breast cancer subclasses.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-02-09.
P2-03-05: Attenuation of TGF-beta Signaling Suppresses Premature Senescence in a p21-Dependent Manner and Promotes Oncogenic ras-Mediated Metastatic Transformation in hTERT-Immortalized Basal-Like Human Mammary Epithelial Cells.
Authors: Lin S , Yang J , Elkahloun AG , Bandyopadhyay A , Wang L , Cornell JE , Yeh I-T , Agyin JK , Sun L-Z 
Published: June 2012

Abstract
Triple negative, basal-like breast cancer is highly aggressive and has a poor prognosis. The molecular mechanisms that drive its progression are elusive and no molecular target has been identified for its prevention or treatment. Here, we demonstrate that triple negative human breast cancer cells and clinical samples show an attenuated transforming growth factor-beta (TGF-β) signaling. Therefore, we developed a series of isogenic basal-like human mammary epithelial cells (HMECs) with altered TGF-β sensitivity and different malignancy, resembling a full spectrum of basal-like breast carcinogenesis, and determined the molecular mechanisms that contribute to oncogene-induced transformation of basal-like HMECs when TGF-β signaling is attenuated. We found that expression of a dominant-negative RII (DNRII) receptor of TGF-β abrogated autocrine TGF-β signaling in telomerase-immortalized HMECs and suppressed H-ras-V12-induced senescence-like growth arrest (SLGA). Furthermore, co-expression of DNRII and H-ras-V12 rendered HMECs to become highly tumorigenic and metastatic in vivo in comparison with H-ras-V12-transformed HMECs that spontaneously escaped H-ras-V12-induced SLGA. By using microarray analysis, we found that p21 is the major player mediating Ras-induce SLGA and attenuated or loss of p21 expression contributed to the escape from SLGA when autocrine TGF-β signaling is blocked in HMECs. Furthermore, knockdown of p21 also suppressed H-ras-V12-induced SLGA. Our results identifies that autocrine TGF-β signaling is an integral part of cellular antitransformation network by suppressing the expression of a host of genes including p21-regulated genes that mediate oncogene-induced transformation in basal-like breast cancer.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-03-05.