bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of integrin as potential molecular markers in human glioblastoma multiforme: Microarray analysis guided by contrast-enhanced MRI
Authors: Yang Y. , Chia Y. , Rubin D. , Harsh G. , Atlas S. , Bednarski M. , Guccione S. 
Published: March 2020

10007 Background: Glioblastoma multiforme (GBM) is a primary brain tumor with poor prognosis and low survival rate. It is characterized as a highly infiltrative and vascular tumor, with morphologic heterogeneity including necrosis regions. We have used contrast enhanced (CE)-MRI to identify regions that differ in vascular permeability as delineated by the small molecule contrast agent Magnevist. Non-necrotic areas with and without contrast enhancement were sampled in the OR. The expression profiles for integrin family genes were investigated and validated using IHC. Genes encoding for integrin subunits with high expression levels in these regions of the tumor were identified and are presented here. Methods: Patients diagnosed with GBM, without any prior procedures were scanned on a GE 1.5T MRI scanner using standard T1- and T2-weighted pulse sequences and Gd(DTPA) as contrast agent. Samples from regions with vascular permeability to the contrast agent (contrast-enhancing, CE) and vascular regions that are not permeable to the contrast agent (non-enhancing, NE) were collected for gene expression profiling using the Affymatrix oligonucleotide microarray system. Genomic data was normalized at probe-level, and analyzed using a modified t-test, Significance Analysis of Microarray (SAM). Significant expression levels were identified. Immunohistochemical staining was used to validate protein expression of selected genes of interest. Results: Tissue samples from the CE and NE regions of 13 patients reveal significantly distinct gene expression patterns. In specific, genes in the integrin family such as integrin αIIb, αV and β3 show high expression levels in the CE region. Immunohistochemical staining confirmed correlation of protein expression patterns with the observed genomic profile. Conclusions: Combination of CE-MRI using the clinical contrast agent and microarray analysis can reveal spatial genomic profiles correlated with imaging features associated with increased vascular permeability, vessel density, and areas of fluid accumulation. The identified genes from the integrin family can be potential molecular targets, and can be used for molecular imaging and therapeutic purposes. No significant financial relationships to disclose.
Microsatellite instability but not thymidylate synthase is a prognostic variable in primary colon cancers from patients treated in 5-FU-based adjuvant studies
Authors: Sinicrope F. A. , Rego R. L. , Halling K. C. , Foster N. R. , Sargent D. J. , Laplant B. , French A. J. , Allegra C. J. , Laurie J. A. , Goldberg R. M. , Thibodeau S. N. 
Published: March 2020

10019 Background: Colon cancers with microsatellite instability (MSI) display resistance to 5-fluorouracil (5-FU), and in vitro resistance can be reversed by restoring DNA mismatch repair proficiency. 5-FU inhibits the thymidylate synthase (TS) enzyme and TS may predict clinical outcome after 5-FU-based chemotherapy. To define molecular predictors of prognosis, we analyzed TS, p53, chromosome 17p allelic imbalance (AI), and patient survival stratified by MSI status. Methods: Primary colon carcinomas from patients enrolled in five 5-FU-based adjuvant therapy trials were analyzed for MSI and 17p AI using 11 microsatellite markers (MSI-H: ≥ 30% of the loci demonstrating instability). For 17p AI, markers included D17S261 and TP53 at or near the p53 locus. Expression of DNA mismatch repair (hMLH1, PharMingen; hMSH2; Oncogene), TS (TS106, Zymed), and p53 (D07, Novacastra) proteins were analyzed by immunohistochemistry. Correlations between markers and associations with overall survival (OS) were determined. Patients were censored at 5 years for DFS and at 8 years post study randomization for overall survival (OS) data. Results: Of 320 Dukes’ stage B2 and C cancers studied, 60 of 320 (19%) were MSI-H. TS expression variables (intensity, extent, weighted score) were similar in MSI-H and MSI stable/low frequency (MSS/MSI-L) cancers; similar results were found using DNA mismatch repair (dMMR) proteins. MSI-H tumors had lower stage (p= 0.0007), fewer metastatic lymph nodes (p= 0.004), and improved OS (vs. MSS/MSI-L tumors; p= 0.01). Loss of dMMR proteins was also associated with better OS (p= 0.006). None of the TS variables were prognostic for OS. Histologic grade (p= 0.0008) and nodal status (p= 0.0002) were associated with OS in contrast to 17p LOH or p53. Only MSI status or dMMR, histologic grade, and tumor stage were independent markers for OS. Conclusions: MSI-H tumors show earlier stage at presentation and better stage-adjusted survival rates. MSI status and TS expression were unrelated and TS was not prognostic, suggesting that TS levels cannot explain therapeutic resistance to 5-FU reported in MSI-H colon cancers. [Table: see text]
Determination of microvessel density by real-time PCR in esophageal cancer: Correlation with histological methods, angiogenic growth factor expression and lymph node metastasis
Authors: Loges S. , Claussen H. , Reichelt U. , Bubenheim M. , Erbersdobler A. , Schurr P. , Yekebas E. , Izbicki J. , Bokemeyer C. , Fiedler W. 
Published: March 2020

10053 Background: Neoangiogenesis and lymphangiogenesis represent prognostic factors in human malignancies. Determination of microvessel density (MVD) by immunohistology is labor-intensive and subject to interobserver variability. We evaluated real time PCR to quantify MVD in primary tumor samples from patients with esophageal cancer. Methods: We performed real-time PCR analyzes of endothel-specific (VE-Cadherin, P1H12, VEGFR-2, tie-2), lymphendothel-specific (Prox, LYVE, VEGFR-3) antigens and of angiogenic growth factors (VEGF-A, VEGF-C, VEGF-D, Ang-1, Ang-2) in primary esophageal carcinoma tissue of 54 patients. These results were compared to MVD determined immunohistochemically by CD31 staining. Results: For validation, MNC samples spiked with HUVECs were analyzed by qPCR for VE-CAD and P1H12 yielding a linear correlation (r=0.99 and 0.96 respectively). Expression of endothelial markers was highly correlated in tumor samples, e.g. CD144 with CD146 τb=0.451, CD144 with VEGFR-2 τb =0.744 and CD144 with tie-2 τb=0.684 (p for all comparisons < 0.0001). QPCR results were compared to MVD determination by CD31 staining in a subgroup of 33 patients. The highest association between both methods was found for CD144 (τb=0.258, p=0.0379) and VEGFR-2 (τb=0.222, p=0.0745) indicating that immunohistology and qPCR yield comparable results. MVD was significantly linked to the expression of VEGF-A, -C,-D and Ang1 and Ang2 (p for all comparisons <0.0001). We analyzed expression of lymphendothelial cell antigens Prox, LYVE and VEGFR-3 for quantification of lymphatic vessels. A close association between the expression of different lymphendothelial factors was seen (LYVE with Prox τb=0.334, p=0.0021, LYVE with VEGFR-3 τb=0.450, p=0.0150). MVD and lymphvessel density was not linked. Lymph node metastases detected on surgical specimen were associated with MVD determined immunohistologically (p=0.003)or by qPCR (p=0.048) and to VEGF-C expression (p=0.04). Conclusions: QPCR analysis of CD144 and VEGFR2 represents a novel tool for quantification of MVD in tumor samples. Expression of VEGFR2 and VEGF-C is associated with lymph node metastasis in patients with esophageal carcinoma. No significant financial relationships to disclose.
A new genomic approach to describe the natural history of node negative breast cancer patients
Authors: Saghatchian M. , Lidereau R. , Delaloge S. , Koscielny S. , Kauffmann A. , Michiels S. , Mathieu M. , Brain E. , Dessen P. , Lazar V. , Tursz T. 
Published: March 2020

10058 Background: Breast cancer is a heterogeneous and complex disease beyond good/bad prognosis groups. Previous studies identified prognostic tools that are undergoing inter-group validation. Our present study aims at better defining pre-malignant vs. malignant lesions and tumors that will give local vs. distant metastases (DM) or early vs. late DM. Methods: We selected 3 groups of patients (pts) in a consecutive series of node-negative breast cancers with a very long follow-up (>10 years (y)) and available frozen tumor samples. Pts who did not receive any adjuvant treatment were selected: 60 with no relapse after 10 y, 29 with early DM before 5 y and 18 with late DM after 5y. Genomic profiling on 44K Agilent pangenomic arrays taking as reference non-relapsing pts (after 10y) or normal breast (Clonetech) allowed us to select specific genes related to DM occurrence or tumorigenesis. Random permutations were performed to assess the statistical significance of our prediction accuracy. Results: 1- we identified a 141 gene-profile and related genes distinguishing tumors with early DM from tumors with late DM (71% prediction accuracy). 2- Similarly, we identified a 285-gene signature for late DM vs. no relapse at 10 y (80% prediction accuracy). Genes were further classified according to family clusters allowing description of genes involved in DM occurrence. 3- a total of 435 genes were also significantly over-expressed in at least 90% of 148 tumours (compared to normal breast). Among those, a selection of the most significant genes were analysed for RT-PC expression throughout the various steps of tumour development (normal breast, benign tumors, in situ carcinomas, invasive carcinomas). Among the 15 most upregulated genes listed, 3 unknown genes appear, that are overexpressed in 98% of the tumors with an average fold change > 11 and are currently explored. Discussion: In this unique series of untreated node-negative breast cancers, identification of molecular profiles of early and late DM could be useful in better early prediction of breast cancer outcome. Based on genomic analysis, a molecular gene based classification of progression from benign tissue to aggressive tumors allows to dissect pathways towards malignancy and to identify early diagnostic markers or targets for prevention. No significant financial relationships to disclose.
Neuregulin (NRG) expression modulates clinical response to trastuzumab in patients with metastatic breast cancer (MBC)
Authors: De Alava E. , Abad M. , Rodriguez C. A. , Montero J. C. , Serrano E. , Ocana A. , Torres A. , Ruiz M. , Cruz J. J. , Pandiella A. 
Published: March 2020

10069 Background: ErbB2 overexpression is the major determinant of response to trastuzumab (Herceptin, H). Recently, some authors have reported the potential role of the expression of NRG on ErbB2 activation in breast cancer cells, and its consequences in response to treatment with H. In the present study we analyze the relationship between response to H and expression of ErbB2 and NRG in patients (pt) with MBC. Methods: Data and frozen tissue samples from 30 consecutive pt with MBC and positive ErbB2 at diagnosis [Immunohistochemistry (IHC)] were collected. All pt were treated with an Herceptin-based regimen. Central Pathologic review of ErbB2 expression was performed in all samples (Herceptest and FISH). NRG expression was studied by IHC on paraffin embedded material and by Western blotting on frozen tissue using an antibody raised against the intracellular domain of NRG. NRG expression by IHC was evaluated using a semiquantitative scoring system that assesses both extension and intensity of cytoplasmic staining. Correlation between ErbB2 and NRG expression and its influence in response to H was analyzed. Characteristics of pt: Median Age: 54; Median mts sites: 2; Treatment: H alone:1 pt; H+Taxol:15 pt; H+Taxol+Carbo: 4 pt; H+Taxol+Lip.Doxorub:2 pt, H+Vinorelbine:7 pt; H+CDDP:1 pt. Results: ORR: 76% (CR:23%, PR:53%). Median TTP: 7.8 m. After central Pathologic review, a total of 20 tumors showed ErbB2 amplification by FISH and 19 of them were Herceptest 3+. A positive correlation was observed between ErbB2 amplification and NRG expression. All complete responses (n=7) were seen in pt with ErbB2 amplification. Interestingly, in the group of tumors without ErbB2 amplification (n=10), 7 had high NRG expression levels, as assessed by IHC. Six out of these 7 tumors having high levels of NRG expression, exhibited partial responses. NRG expression, in turn, did not have impact on response among ErbB2 amplified tumors. Conclusions: These preliminary results suggest that responses to H regimens in pt with MBC can be seen in pt lacking ErbB2 amplification in presence of high levels of expression of transmembrane ligand NRG. This suggests that the group of pt that may benefit from treatment with H could be broader than currently established. A confirmatory study is ongoing in a larger series. No significant financial relationships to disclose.
Immunization with MUC1/X protein enhances cDNA immunization in generating anti-MUC1 alpha/beta junction antibodies that target cancer cells
Authors: Rubinstein D. B. , Ziv R. , Karmely M. , Leitner O. , Wreschner D. 
Published: March 2020

10072 Background: MUC1, a glycoprotein highly expressed in epithelial malignancies including breast, prostate, and ovarian, and on the malignant cells of multiple myeloma has generated considerable interest as a tumor marker and target for tumor killing. The most intensively studied MUC1 protein is a type I transmembrane protein (MUC1/TM) which is proteolytically cleaved soon after synthesis into α and β subunits which bind in a strong non-covalent interaction. Almost all antibodies generated to date against MUC1 recognize epitopes within the highly immunogenic tandem-repeat-array. A major shortcoming in use of such antibodies is the fact that the tandem-repeat-array-containing part of MUC1 is shed from the cell surface into the circulation. Soluble, shed MUC1 sequesters circulating anti-tandem-repeat-array antibodies, limiting their ability to reach targeted MUC1-expressing cells. Antibodies to MUC1 epitopes tethered to the cell surface would likely be more effective therapeutic agents. Despite efforts in recent years, such antibodies have remained elusive; generation of anti-cell antibodies requires characterization of cell-bound epitopes. The junction of the MUC1 α-subunit binding the membrane-tethered β-subunit provides such an epitope. Methods: By use of a novel protocol, entailing immunization with MUC1/TM cDNA and boosting with MUC1/X protein, a MUC1 isoform lacking the tandem-repeat-array, we generated monoclonal antibodies designated DMC209 which recognize the MUC1 α/β junction. Results: DMC209 is exquisitely unique for the target site; all amino acid mutations which abrogate MUC1 cleavage also abrogate DMC209 binding. Additionally, DMC209 binds the MUC1 α/β junction on cell-tethered tandem-repeat-array-containing MUC1 (MUC1/TM) on breast and ovarian cancer, and on myeloma cells. Conclusion: DMC209 is likely to have clinical application by targeting MUC1-expressing cells directly, and as an immunotoxin conjugate. Moreover, the novel high-titer immunization procedure used in generating DMC209 can be used to generate anti-MUC1 α/β junction antibodies acting as ligand and, analogously to herceptin, may have direct cytotoxic activity. No significant financial relationships to disclose.
Predicting survival and metastasis in soft tissue sarcoma: A gene microarray study using RNA derived from archival paraffin-embedded tumours
Authors: Linton K. M. , Hey Y. , Jeziorska M. , Wilson C. L. , Miller C. , Saunders E. , Pepper S. , Freemont A. , Radford J. A. 
Published: March 2020

10074 Background: The genetic determinants of survival and metastasis in soft tissue sarcoma (STS) are poorly understood and progress in this field has been limited by the rare nature of STS and the need for fresh/frozen tissue (FT) for gene microarray analyses. Our objective was to determine whether valid gene microarray data can be obtained from archival formalin-fixed paraffin-embedded tumours (FFPET) in order to retrospectively identify prognostic STS gene signatures and potential molecular targets from patients with known clinical outcome data. Methods: Total RNA was extracted from macrodissected viable tumour tissue for 34 FFPET (14 liposarcomas, 11 leiomyosarcomas and 9 synovial sarcomas) and two paired FFPET and FT primary leiomyosarcomas (modified Optimum kit, Ambion, for FFPET RNA extraction and Trizol, Invitrogen, for FT RNA extraction). One-cycle labelled cRNA was hybridised to Affymetrix U133 Plus 2 microarrays, and strict QC protocols used to identify 19 arrays for further analysis using R and BioConductor. Log rank regression and Kaplan-Meier plots of disease-specific survival were performed to identify genes predictive of survival and/or metastasis. Results: Similar fold changes in gene expression were obtained for paired cRNA samples, although the number of genes called present was lower for cRNA from FFPET. Five hundred genes (selected from FFPET cRNA arrays) were discriminatory for survival (p=0.0006 to 0.0088) across all STS subtypes and correctly assigned 17/19 cases by leave-one-out cross validation. Many are known tumour prognostic genes, significant for survival, metastasis, invasion, angiogenesis and apoptosis. Twenty-five novel and known candidate genes (with 3–10 fold differential expression) were selected for validation on 19 test and 60 independent cases by RT-PCR for relative gene expression and immunohistochemistry for protein detection. Conclusions: We have shown that prognostic information can be derived from archival FFPET, permitting the identification of candidate prognostic genes and therapeutic targets in STS, and opening the way for studies in other tumours where FT is unavailable. No significant financial relationships to disclose.
Estrogen receptor β represses breast tumor growth and angiogenesis in vivo
Authors: Hartman J. , Lindberg K. , Inzunza J. , Wan J. , Ström A. , Gustafsson J. 
Published: March 2020

10101 Background: Estrogens are well known stimulators of breast cancer cell growth in vitro as well as in vivo. Two different estrogen receptors exist, namely estrogen receptor (ER) α and β. ERα mediates the proliferative effect of estrogen in breast cancer cells and we have earlier shown that ERβ inhibits cell-cycle progression in vitro. Estrogens are well known stimulators of in vivo breast cancer cell growth as well as angiogenesis, and the effect is mediated through ERα. The function of ERβ in this context is not well understood. Methods: We have used ERα-positive T47D breast cancer cells stably transfected with a Tet/Off regulated ERβ expression vector system. The ERβ-inducible tumor cells are studied in vitro as well as in vivo. Results: By transplanting ERβ-inducible breast cancer cells into SCID-mice, we show that ERβ inhibits tumor growth and reduces the volume of established tumors. Furthermore, we show by immunohistochemistry, that the number of blood microvessels in the tumor periphery is decreased by ERβ expression, counteracting the well-known pro-angiogenic effect of ERα. By Western blot analysis on tumor extracts, we show that the concentration of the important pro-angiogenic growth factors VEGF and bFGF, normally expressed by breast tumor cells, is decreased in the ERβ-expressing tumors compared to the normal tumors. To exclude that the observed anti-angiogenic effect is just a result of reduced tumor growth, we incubated Tet/Off regulated ERβ expressing cells in vitro, during non-hypoxic conditions. We found that the expression of ERβ leads to decreased expression of VEGF and PDGFβ at the mRNA and protein-levels. In transient transfection assays, we found estrogen-ERα mediated up regulation of VEGF, PDGFβ and bFGF-promoter activities in T47D cells, and these activities were all suppressed following co-transfection with an ERβ-expression vector. Conclusions: We conclude that ERβ inhibits growth factor expression at transcriptional level in breast cancer cells; taken together, our data indicates that ERβ inhibits growth and angiogenesis of tumors formed by T47D breast cancer cells. This makes ERβ an interesting therapeutic target in breast cancer and perhaps treatment with the newly designed ERβ-selective ligands might work as a new anti-proliferative and anti-angiogenic therapy. No significant financial relationships to disclose.
Heterogeneity of epidermal growth factor receptor (EGFR) expression and variation in immunohistochemistry (IHC) testing may affect access to EGFR-targeted therapy in patients with advanced colorectal cancer (CRC)
Authors: Beer M. W. , Ung C. , Bacus S. S. , McCollum A. D. , Ervin-Haynes A. , Schinagl R. , Youssoufian H. , Rowinsky E. 
Published: March 2020

10104 Introduction: Cetuximab, an IgG1 MAb directed at the EGFR, has demonstrated benefit in pts with CRC and is currently indicated in EGFR expressing CRC. However, previous reports show heterogeneous EGFR expression between anatomic sites and suggest that results from IHC testing vary between labs. To assess this variability and the resultant impact on pt access to EGFR-targeted therapy, we compared EGFR results reported to clinicians by hospital and reference labs with those obtained at a single central lab. Methods: Formalin-fixed, paraffin-embeded CRC specimens submitted through the Targeted Diagnostics Advocacy Program (tdap) were assayed using the Dako EGFR pharmDx kit and scored using the FDA-approved package insert. If the initial specimen tested EGFR negative (−), additional specimens were requested for assay. Prior EGFR results, lab, and specimen ID were documented through review of anatomic pathology reports. Results: Of 332 evaluable CRC pts tested, 245 pts (74%) were previously reported as EGFR(−). After retesting, 167 (68%) pts were found to be EGFR The originally tested tumor block was retested in 82 pts and 41 pts (50%) were found to be EGFR (+). Retest results were similar for both hospital and reference labs. Of 66 pts with >1 specimen from the same anatomic site tested, 21 (32%) were discordant (at least 1 (+) and one (−) result). Results from different anatomic sites were compared for 65 pts, 34 (52%) of which were discordant (+). Overall, 231 pts (70%) tested EGFR(+). Conclusion: This study shows marked heterogeneity in EGFR expression by IHC both within and across anatomic sites from the same pt as well as variability in both hospital and reference pathology labs. This variability undoubtedly has an adverse impact on pt access to EGFR-targeted therapy. The impact of EGFR heterogeneity may be reduced through the assay of multiple specimens. [Table: see text] [Table: see text]