bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential modulation of a polymorphism in the COOH terminus of the α-subunit of the human epithelial sodium channel by protein kinase Cδ
Authors: Yan Wusheng , Suaud Laurence , Kleyman Thomas R. , Rubenstein Ronald C.
Published: September 2005

The A663T polymorphism of the α-subunit of the human epithelial sodium channel (hENaC) increases the functional and surface expression of αβγ-hENaC in Xenopus laevis oocytes. The context of this residue in the COOH terminus of α-hENaC is important for this effect, as a homologous change in murine ENaC (mENaC), A692T, does not alter functional and surface expression of mENaC. Query of a phosphoprotein database suggested that the α-T663 residue might be phosphorylated by PKCδ. General inhibition of PKC with calphostin C decreased the functional and surface expression of αT663-hENaC and not αA663-hENaC, and was without effect on αA692-mENaC, αT692-mENaC, and a chimeric m(1–678)/h(650–669)αT663, mβγ-ENaC. These data suggest that residues outside of the α-hENaC COOH terminus are important for modulation of αT663-hENaC trafficking by PKC. In contrast, expression of PKCδ decreased the functional and surface expression of αT663-hENaC and the functional expression of m(1–678)/h(650–669)αT663, mβγ-ENaC, and was without effect on αA663-hENaC, αA692-mENaC, or αT692-mENaC. PKCδ did not phosphorylate the COOH terminus of either αT663-hENaC or αA663-hENaC in vitro, suggesting that it acts indirectly to regulate hENaC trafficking. αT663-hENaC was retrieved from the oocyte membrane more slowly than αA663-hENaC, and calphostin C increased the rate of αT663-hENaC removal from the oocyte membrane to a rate similar to that of αA663-hENaC. In contrast, PKCδ did not alter the rate of removal of αT663-hENaC from the oocyte membrane, suggesting that PKCδ altered rates of αT663-hENaC biosynthesis and/or delivery to the plasma membrane. These data are consistent with PKC isoform-specific effects on the intracellular trafficking of αT663- vs. αA663-hENaC.
Combined 5-Fluorouracil/Systemic Interferon-β Gene Therapy Results in Long-Term Survival in Mice with Established Colorectal Liver Metastases
Authors: Choi Eugene A. , Lei Hanqin , Maron David J. , Mick Rosemarie , Barsoum James , Yu Qian-chun , Fraker Douglas L. , Wilson James M. , Spitz Francis R.
Published: September 2005

Preclinical in vitro and in vivo studies have demonstrated synergistic interactions between 5-fluorouracil (5-FU) and type I and II IFNs against human colorectal cancer cells. Despite these activities, randomized human trials have failed to identify a clinical benefit for this combination treatment. These limited clinical results may be secondary to the short half-life of recombinant IFN protein and the increased systemic toxicities of 5-FU/IFN combinations. We have previously reported an adenoviral-mediated IFN-β gene therapy strategy, which may circumvent the pitfalls of recombinant IFN therapy. However, a dose-dependent toxicity and acute inflammatory response to systemically administered adenovirus vectors may limit the clinical application of this therapy. The combination of adenoviral-mediated IFN-β gene therapy and 5-FU resulted in tumor regression, apoptosis, and improved survival in an established liver metastases model. These therapeutic effects were observed at a significantly lower vector dose than we had previously reported and with limited toxicity. This approach may allow for an effective clinical application of this therapy and warrants additional investigation.
Clinical Significance of Immunocytochemical Detection of Tumor Cells Using Digital Microscopy in Peripheral Blood and Bone Marrow of Breast Cancer Patients
Authors: Pierga Jean-Yves , Bonneton Charlyne , Vincent-Salomon Anne , de Cremoux Patricia , Nos Claude , Blin Nathalie , Pouillart Pierre , Thiery Jean-Paul , Magdelénat Henri
Published: September 2005

Purpose: The presence of tumor cells in bone marrow has been reported to represent an important prognostic indicator in breast cancer, but the clinical significance of circulating cells in peripheral blood is less well known. The aim of this study was to evaluate the feasibility of identifying cytokeratin (CK)-expressing cells in peripheral blood with an automat-assisted immunohistochemical detection system and to compare it with detection of tumor cells in bone marrow samples.

Experimental Design: Cytospun Ficoll fractions of peripheral blood and bone marrow were obtained simultaneously in 114 breast cancer patients at different stages of the disease (I to IV) before treatment with chemotherapy. The pancytokeratin (CK) monoclonal antibody A45-B/B3 (anti-CKs 8, 18, and 19) was used for epithelial cell detection. Immunostained cells were detected by an automated cellular imaging system (ChromaVision Medical System).

Results: CK+ cells were detected in 28 (24.5%) patients in blood and in 67 (59%) patients in bone marrow. Twenty-six (93%) patients with CK-positive cells in blood also had positive bone marrow (P < 0.001). Positive cells were detected in peripheral blood in 3/39 (7.5%) operable breast cancers (stage I/II), 9 of 36 (25%) locally advanced breast cancers (stage III), and 16 of 39 (41%) patients with metastatic disease (stage IV; P = 0.017). In the subgroup of nonmetastatic patients (n = 75), prognostic factors for poor disease-free survival were: absence of estrogen receptor; presence of CK+ cells in bone marrow (P = 0.012); clinical nodal involvement; large tumor size (T4); and presence of tumor emboli. Presence of circulating CK+ cells in the peripheral blood was not statistically correlated with disease-free survival. On multivariate analysis, independent indicators for disease-free survival were: absence of estrogen receptor (P = 0.043) and presence of CK+ cells in bone marrow (P = 0.076).

Conclusions: The clinical relevance of circulating epithelial cells as a prognostic factor is not supported by the present data, especially in comparison with tumor cells in the bone marrow. However, this method of detection may be useful to monitor the efficacy of treatment in advanced or metastatic breast cancer.
Prognostic Value of CD40 in Adult Soft Tissue Sarcomas
Authors: Ottaiano Alessandro , De Chiara Anna , Perrone Francesco , Botti Gerardo , Fazioli Flavio , De Rosa Vincenzo , Mozzillo Nicola , Ravo Vincenzo , Morrica Brunello , Gallo Ciro , Pisano Carmela , Napolitano Maria , Ascierto Paolo Antonio , Iaffaioli Rosario Vincenzo , Apice Gaetano
Published: September 2005

Purpose: The purpose is to evaluate the expression of CD40, a membrane protein predominantly expressed on B cells, dendritic cells, and macrophages, in a series of adult soft tissue sarcomas and to test its possible prognostic value.

Experimental Design: CD40 expression was studied by immunohistochemistry. Correlations with other baseline characteristics of patients and tumors were analyzed with χ2 test. The prognostic value was studied with univariable and multivariable analysis adjusted by age, sex, tumor size, grade, location, and distant metastases.

Results: Eighty-two patients, between January 1994 and May 2001, were analyzed. Membrane or cytoplasmic staining for CD40 protein was absent in 30% of the tumors but present in <10% of cells in 22 (27%), in 10% to 50% in 23 (28%), and in >50% of cells in 12 (15%) tumors. There was no correlation between CD40 expression and age, sex, size, grade, and location of the primary tumor and distant metastases. With 61 patients (74.4%) progressed and 31 (37.8%) dead, CD40 expression was a significant prognostic factor for disease-free and overall survival at univariable and multivariable analysis. Patients with tumors expressing CD40 in >50% of cells had a dramatically unfavorable prognosis with median disease-free and overall survival of 7 and 17 months, respectively, and hazard ratios of relapse and death as compared with patients with CD40-negative tumors of 2.89 (95% confidence interval: 1.26–6.60) and 6.92 (95% confidence interval: 2.18–22.0), respectively.

Conclusions: These data suggest that expression of CD40 protein in >50% of cells might indicate an unfavorable prognosis in adult soft tissue sarcomas.
Quantitative
TP73
Transcript Analysis in Hepatocellular Carcinomas
Authors: Stiewe Thorsten , Tuve Sebastian , Peter Martin , Tannapfel Andrea , Elmaagacli Ahmet H. , Pützer Brigitte M.
Published: September 2005

Purpose: The p53 family member p73 displays significant homology to p53, but data from primary tumors demonstrating increased expression levels of p73 in the absence of any gene mutations argue against a classical tumor suppressor function. A detailed analysis of the p73 protein in tumor tissues has revealed expression of two classes of p73 isoforms. Whereas the proapoptotic, full-length, transactivation-competent p73 protein (TA-p73) has a putative tumor suppressor activity similar to p53, the antiapoptotic, NH2-terminally truncated, transactivation-deficient p73 protein (ΔTA-p73) has been shown to possess oncogenic activity. The oncogenic proteins can be generated by the following two different mechanisms: (a) aberrant splicing (p73Δex2, p73Δex2/3, ΔN′-p73) and (b) alternative promoter usage of a second intronic promoter (ΔN-p73). The purpose of our study was to elucidate the origin of ΔTA-p73 isoforms in hepatocellular carcinomas.

Experimental Design: We analyzed the underlying mechanisms of p73 overexpression in cancer cells by quantification of p73 transcripts from 10 hepatocellular carcinoma patients using isoform-specific real-time reverse transcription-PCR.

Results: Our data demonstrate that only aberrantly spliced ΔTA-p73 transcripts from the TA promoter show significantly increased expression levels in the tumor whereas the ΔN-p73 transcript generated from the second promoter is not significantly up-regulated.

Conclusions: Although we only analyzed 10 patient samples the results strongly suggest that the elevated activity of the first promoter (TA promoter) accounts for high-level expression of both full-length TA-p73 and aberrantly spliced ΔTA-p73 isoforms in hepatocellular carcinoma tissues.
Phase I and Pharmacokinetic Study of the Polyamine Synthesis Inhibitor SAM486A in Combination with 5-Fluorouracil/ Leucovorin in Metastatic Colorectal Cancer
Authors: van Zuylen Lia , Bridgewater John , Sparreboom Alex , Eskens Ferry A. L. M. , de Bruijn Peter , Sklenar Ivo , Planting Andre S. T. , Choi Les , Bootle Douglas , Mueller Christian , Ledermann Jonathan A. , Verweij Jaap
Published: September 2005

Purpose: The purpose of our study was to determine the maximum-tolerated dose, dose-limiting toxicity, safety profile, and pharmacokinetics of the polyamine synthesis inhibitor SAM486A given in combination with 5-fluorouracil/leucovorin (5-FU/LV) in cancer patients.

Experimental Design: Patients with advanced colorectal cancer were treated with 5-FU [bolus (400 mg/m2) followed by a 22-h infusion (600 mg/m2)] and LV (200 mg/m2) and escalating doses of SAM486A, 1–3-h infusion daily for 3 days. Plasma sampling was performed to characterize the pharmacokinetics and pharmacodynamics of the combination

Results: Twenty-seven patients with metastatic colorectal cancer and 1 with pseudomyxoma peritonei were treated. Twenty-six patients received SAM486A in the combination at doses ranging from 25 to 150 mg/m2/day. Dose-limiting toxicity consisting of fatigue grade 3 was seen at 150 mg/m2/day. Other adverse events included neutropenia, hand and foot syndrome, nausea, vomiting, diarrhea, and constipation. Fifteen of 26 patients evaluable for best response according to the Southwest Oncology Group criteria achieved a partial response [8 (30%) of 26] or stable disease [9 (35%) of 26]. SAM486A did not influence the pharmacokinetics of 5-FU, and SAM486A clearance was similar to that when used as a single agent.

Conclusions: The novel molecular agent SAM486A is tolerable and safe in combination with a standard 5-FU regimen in patients with advanced colorectal cancer. The dose of SAM486A recommended for additional studies with this combination is 125 mg/m2/day. A disease-directed evaluation of SAM486A using this regimen is warranted.
Expression of Trypsinogen-1, Trypsinogen-2, and Tumor-Associated Trypsin Inhibitor in Ovarian Cancer
Authors: Paju Annukka , Vartiainen Juhani , Haglund Caj , Itkonen Outi , von Boguslawski Kristina , Leminen Arto , Wahlström Torsten , Stenman Ulf-Håkan
Published: September 2005

Purpose: The purpose is to study the prognostic significance of tissue expression of trypsinogen-1, trypsinogen-2, and tumor-associated trypsin inhibitor (TATI) and serum concentration of trypsinogen-2, trypsin-2-API (complex of trypsin-2 with α-1-proteinase inhibitor), and TATI in epithelial ovarian cancer.

Experimental Design: Expression of trypsinogen-1, trypsinogen-2, and TATI was determined by immunohistochemistry with monoclonal antibodies in tissue sections of tumors from 119 patients with untreated primary epithelial ovarian cancer. Preoperative serum concentrations of trypsinogen-2, trypsin-2-API and TATI were analyzed using specific immunoassays.

Results: Fifty-four percent of the tumors expressed trypsinogen-1, 45% expressed trypsinogen-2, and 30% expressed TATI. In patients with stage III and IV disease, TATI tissue expression (P = 0.002) and elevated TATI concentration in serum (P = 0.048) were associated with adverse cancer-specific and progression-free survival in univariate analysis. In multivariate analysis, TATI tissue expression (P = 0.005), tumor grade (P = 0.0001), histological type (P = 0.02), and stage (P = 0.0005) were independent prognostic factors for adverse cancer-specific survival and TATI tissue expression (P = 0.006) and grade (P = 0.0003) for progression-free survival. In multivariate analysis of all patients and those with advanced disease, serum trypsin-2-API concentration was an adverse prognostic factor for cancer-specific and progression-free survival, and it was independent of stage and histological type of the tumor (P ≤ 0.01).

Conclusions: Tissue expression of TATI and an elevated preoperative serum concentration of trypsin-2-API are strong independent prognostic factors in advanced epithelial ovarian cancer. These results suggest that trypsin expression plays a role in the progression of ovarian cancer. TATI and trypsin-2-API are of potential use as an aid for stratification of randomized studies and for selecting treatment strategies.
Phase II Study of SU5416, a Small Molecule Vascular Endothelial Growth Factor Tyrosine Kinase Receptor Inhibitor, in Patients with Refractory Multiple Myeloma
Authors: Zangari Maurizio , Anaissie Elias , Stopeck Alison , Morimoto Alyssa , Tan Nguyen , Lancet Jeffrey , Cooper Maureen , Hannah Alison , Garcia-Manero Guillermo , Faderl Stephan , Kantarjian Hagop , Cherrington Julie , Albitar Maher , Giles Francis J.
Published: September 2005

Purpose: Increased bone marrow angiogenesis and vascular endothelial growth factor (VEGF) levels are of adverse prognostic significance in patients with multiple myeloma (MM). VEGF, a soluble circulating angiogenic molecule, acts via receptor tyrosine kinases, including VEGF receptor 2. SU5416 is a small molecule VEGF receptor 2 inhibitor.

Experimental Design: Adult patients with advanced MM were entered on a multicenter phase II study.

Results: Twenty-seven patients (median age 69, range 39–79), median 4 (0–10) lines of prior therapy, 14 with prior thalidomide therapy, received SU5416 at 145 mg/m2 twice weekly i.v. for a median of two 4-week cycles (range 0.2–9). Grade 3/4 toxicities were rarely observed; the most frequent was thrombocytopenia (12%). Mild-to-moderate toxicities included nausea (63%), headache (56%), diarrhea, vomiting (both 37%), and fatigue (33%). There were three thromboembolic episodes and five cases of new onset hypertension. Two (7%) patients did not complete the first 4-week cycle of therapy because of adverse events (pneumonia and headache). There were no objective responses. Four patients had disease stabilization for ≥4 months. A decrease in median VEGF plasma levels was observed in patients with stable disease (n = 7) compared with patients with progressive disease (n = 5). Overall median survival was 42 weeks (range 3–92+).

Conclusions: Although SU5416 had minimal clinical activity, signs of biological activity (decrease in plasma VEGF levels) suggest that angiogenic modulation may be of value in patients with MM.
Detection and Grading of Soft Tissue Sarcomas of the Extremities with 18F-3′-Fluoro-3′-Deoxy-l-Thymidine
Authors: Cobben David C. P. , Elsinga Philip H. , Suurmeijer Albert J. H. , Vaalburg Wim , Maas Bram , Jager Piet L. , Hoekstra Harald J.
Published: September 2005

Purpose: The aim of the study was to investigate the feasibility of 18F-3′-fluoro-3′-deoxy-l-thymidine positron emission tomography (FLT-PET) for the detection and grading of soft tissue sarcoma (STS).

Experimental Design: Nineteen patients with 20 STSs of the extremities were scanned, using attenuation corrected whole-body FLT-PET. Standardized uptake values (SUVs) and tumor:nontumor ratios (TNTs) were compared with histopathological parameters using French and Japanese grading systems.

Results: Mean SUV, maximal SUV, and TNT could differentiate between low-grade (grade 1; n = 6) STS and high-grade (grade 2 and 3; n = 14) STS according to the French grading system (P = 0.001). Mean SUV, max SUV, and TNT correlated with mitotic score, MIB-1 score, the French and Japanese grading system (• = 0.550–0.747).

Conclusions: FLT-PET is able to visualize STS and differentiate between low-grade and high-grade STS. The uptake of FLT correlates with the proliferation of STS.
Predicting Early Failure after Adjuvant Chemotherapy in High-Risk Breast Cancer Patients with Extensive Lymph Node Involvement
Authors: Faneyte Ian F. , Peterse Johannes L. , van Tinteren Harm , Pronk Corina , de Vries Elisabeth G. E. , Rodenhuis Sjoerd , van de Vijver Marc J.
Published: September 2005

Purpose: There is limited knowledge of risk factors for breast cancer recurrence within 2 years. This study aimed to predict early failure and identify high-risk patients for prognostic and therapeutic purposes.

Experimental Design: We studied 739 patients from a randomized trial who were <56 years of age and had ≥4 or more positive lymph nodes, no distant metastases, and no previous other malignancies. After complete surgical treatment, patients received conventional-dose anthracycline-based chemotherapy or a high-dose scheme of anthracycline-based plus alkylating chemotherapy. We assessed clinical and (immuno)histological parameters to predict recurrence within 2 years.

Results: Early failure occurred in 19% (n = 137). Median survival after early failure was limited to 0.7 year. Estrogen and progesterone receptor negativity and visceral relapse predicted poor prognosis. Early failure was associated with young age, large tumors, high histological grade, angio-invasion, apical node metastasis, and ≥10 involved nodes. Estrogen receptor, progesterone receptor, and p27 negativity; HER2 overexpression; and p53 positivity also predicted early failure. The surgical or chemotherapy regimen and histological type did not. The same parameters except tumor size were associated with early death. Grade III, ≥10 involved nodes, and estrogen receptor negativity were independently associated with early failure and together identified a subset of patients (7%) with 3-fold increased early failure and 5-fold increased early death.

Conclusions: Early failure is associated with poor survival. The combination of three commonly determined parameters constitutes a strong predictive model for early failure and death.
Generation of Kidney Cancer-Specific Antitumor Immune Responses Using Peripheral Blood Monocytes Transduced With a Recombinant Adenovirus Encoding Carbonic Anhydrase 9
Authors: Mukouyama Hideki , Janzen Nicolette K. , Hernandez Jose M. , Lam John S. , Caliliw Randy , Wang Allen Y. , Figlin Robert A. , Belldegrun Arie S. , Zeng Gang
Published: September 2005

Purpose: Carbonic anhydrase 9 (CA9) is the most promising molecular marker described for renal cell carcinoma (RCC) to date. We investigated whether transduction of monocytes from peripheral blood with adenovirus encoding the CA9 gene (AdV-CA9) could stimulate a T-cell mediated immune response against cancer cells expressing CA9. The ability to consistently generate a T-cell response is an important step toward the development of a CA9-specific RCC vaccine.

Experimental Design: AdV-CA9 was generated using the AdEasy system. AdV-CA9-transduced peripheral blood mononuclear cell (PBMC)-derived monocytes were used to raise CTLs from autologous peripheral blood lymphocytes (PBLs). The ability of CTLs to lyse targets expressing CA9 was assessed by 51Cr-release.

Results: Monocytes were efficiently transduced with AdV-CA9. In five of six experiments, AdV-CA9-transduced monocytes were able to induce a population of CTLs from bulk PBLs. CTLs were capable of lysing autologous, but not allogeneic monocytes expressing CA9. Furthermore, CTLs were able to lyse autologous RCC tumor cells expressing CA9. The ability of CTLs to lyse relevant targets was blocked by anti-CD3, anti-CD8, and anti-MHC class I antibodies demonstrating a MHC class I restricted response.

Conclusions: These results suggest that PBMC-derived monocytes transduced with AdV-CA9 can generate RCC-specific MHC class I restricted CTLs capable of lysing CA9-expressing cancer cells. Transduction of PBMC-derived monocytes with adenovirus provides a simple and effective alternative to the use of dendritic cells for the induction of antigen-specific CTL.
Somatostatin Receptor Subtype 2 Is Expressed by Supratentorial Primitive Neuroectodermal Tumors of Childhood and Can Be Targeted for Somatostatin Receptor Imaging
Authors: Frühwald Michael C. , Rickert Christian H. , O’Dorisio M. Sue , Madsen Mark , Warmuth-Metz Monika , Khanna Geetika , Paulus Werner , Kühl Joachim , Jürgens Heribert , Schneider Peter , Müller Hermann L.
Published: September 2005

Purpose: Although gliomas predominate among central nervous system (CNS) neoplasms in adulthood, embryonal tumors are the most common malignant brain tumors in children. Despite novel treatment approaches, including improved radiotherapy and high-dose chemotherapy, survival rates remain unsatisfactory. The timely diagnosis of residual or recurrent embryonal CNS tumors and thus the earliest possible time point for intervention is often hampered by inaccuracies of conventional imaging techniques. Novel and refined imaging methodologies are urgently needed.

Experimental Design: We have previously demonstrated the use of somatostatin receptor imaging (SRI) in the diagnosis of recurrent and residual medulloblastomas. Here, we evaluated somatostatin receptor type 2 (sst2) expression using an antibody in an array of CNS tumors of childhood. Eight high-grade gliomas, 4 atypical teratoid/rhabdoid tumors, 7 supratentorial primitive neuroectodermal tumors (stPNET), 1 medulloepithelioma (ME), and 8 ependymomas were screened. Tumors positive in vitro were additionally analyzed in vivo using SRI.

Results: Abundant expression of somatostatin receptor type 2 in stPNET, a ME, and ependymomas warranted in vivo imaging of 7 stPNET, 1 rhabdomyosarcoma, 3 ependymomas, 1 ME, and 1 glioblastoma. Although SRI was positive in 6/7 stPNET, 1 rhabdomyosarcoma, and 1 ME, none of the ependymomas nor the glioblastoma could be imaged using SRI. In selected cases SRI was more sensitive in the detection of relapse than conventional imaging by magnetic resonance imaging and computed tomography.

Conclusions: SRI should be considered in the evaluation of residual or recurrent embryonal CNS tumors, especially stPNET. The strengths of SRI lie in the differentiation of reactive tissue changes versus residual or recurrent tumor, the detection of small lesions, and possibly in the distinction of stPNET from gliomas.
Alterations of the Tumor Suppressor Gene
Parkin
in Non-Small Cell Lung Cancer
Authors: Picchio Maria Cristina , Martin Eric Santos , Cesari Rossano , Calin George Adrian , Yendamuri Sai , Kuroki Tamotsu , Pentimalli Francesca , Sarti Manuela , Yoder Kristine , Kaiser Larry R. , Fishel Richard , Croce Carlo Maria
Published: September 2005

Purpose: Parkin, a gene mutated in autosomal recessive juvenile Parkinsonism and mapped to the common fragile site FRA6E on human chromosome 6q25-q27, is associated with a frequent loss of heterozygosity and altered expression in breast and ovarian carcinomas. In addition, homozygous deletions of exon 2 creating deleterious truncations of the Parkin transcript were observed in the lung adenocarcinoma cell lines Calu-3 and H-1573, suggesting that the loss of this locus and the resulting changes in its expression are involved in the development of these tumors.

Experimental Design: We examined 20 paired normal and non-small cell lung cancer samples for the presence of Parkin alterations in the coding sequence and changes in gene expression. We also restored gene expression in the Parkin-deficient lung carcinoma cell line H460 by use of a recombinant lentivirus containing the wild-type Parkin cDNA.

Results: Loss of heterozygosity analysis identified a common region of loss in the Parkin/FRA6E locus with the highest frequency for the intragenic marker D6S1599 (45%), and semi-quantitative reverse transcription-PCR revealed reduced expression in 3 of 9 (33%) lung tumors. Although we did not observe any in vitro changes in cell proliferation or cell cycle, ectopic Parkin expression had the ability to reduce in vivo tumorigenicity in nude mice.

Conclusion: These data suggest that Parkin is a tumor suppressor gene whose inactivation may play an important role in non-small cell lung cancer tumorigenesis.
Epithelial-Cadherin and β-Catenin Expression Changes in Pancreatic Intraepithelial Neoplasia
Authors: Al-Aynati Maamoun M. , Radulovich Nikolina , Riddell Robert H. , Tsao Ming-Sound
Published: September 2005

Purpose: Cadherins and associated catenins are important mediators of epithelial cell-cell adhesion, as well as the Wnt-signaling pathway. Significant changes in their expression or structure have been implicated in malignancy. This study aimed to investigate the epithelial-cadherin (E-cadherin) and β-catenin expression changes during multistage, pancreatic ductal carcinogenesis.

Experimental Design: Ninety-four Whipple resection specimens were retrieved from the surgical pathology files of the University Health Network (Toronto, Canada), from which tissue microarray blocks containing 36 pancreatic ductal adenocarcinomas, 34 PanIN-1A lesions, 28 PanIN-1B lesions, 27 PanIN-2 lesions, 16 PanIN-3 lesions, and 32 normal ducts were constructed. The E-cadherin, β-catenin, and the phosphorylated glycogen synthase kinase-3β of the Wnt/β-catenin pathway were immunohistochemically evaluated in these duct/PanIN lesions.

Results: There was marked increase in the cytoplasmic E-cadherin expression in PanIN lesions (P < 0.0001) and adenocarcinoma (P = 0.005) compared with normal pancreatic ducts. In contrast, reduced/loss of E-cadherin membranous expression was also significant in ductal adenocarcinoma compared with both the PanIN lesions (P < 0.0001) and normal ducts (P = 0.05). The β-catenin expression showed significantly more frequent aberrant nuclear localization in high-grade PanIN lesions, particularly PanIN2 and in adenocarcinoma compared with normal ducts or low grade PanIN lesions (P < 0.0001). However, there was a lack of correlation between phosphoSer9-glycogen synthase kinase-3β cytoplasmic expression and β-catenin aberrant nuclear expression (P = 0.07).

Conclusions: Aberration in the expression of E-cadherin and its associated β-catenin is evident in pre-invasive (PanIN) neoplastic pancreatic duct cells, suggesting involvement of pathways leading to β-catenin stabilization during pancreatic duct cell carcinogenesis.
Inhibition of Orthotopic Human Bladder Tumor Growth by Lentiviral Gene Transfer of Endostatin
Authors: Kikuchi Eiji , Menendez Silvia , Ohori Makoto , Cordon-Cardo Carlos , Kasahara Noriyuki , Bochner Bernard H.
Published: September 2005

Purpose: Inhibitors of endothelial cell proliferation, such as endostatin, result in suppression of tumor-associated angiogenesis and can achieve growth-inhibitory effects depending on the type of tumor treated. The purpose of this study was to investigate whether local overexpression of endostatin could serve to diminish tumor growth of bladder cancer in vivo.

Experimental Design: We examined the capability of lentiviral-mediated gene transfer in vitro and therapeutic effects of lentivirus-based vectors expressing endostatin on tumor growth using an orthotopic human bladder tumor model.

Results: We found that self-inactivating lentivirus vectors containing green fluorescent protein, alone or in combination with endostatin, were capable of efficient and stable gene transfer to a variety of human bladder tumor cell lines. The production and secretion of endostatin from lentivirus-transduced KU-7 human bladder cancer cells was confirmed by Western blot and competitive enzyme immunoassay. Intravesical instillation of untransduced, green fluorescent protein control lentivirus-transduced, and endostatin-transduced KU-7 cells was performed in murine models to establish orthotopic tumors. Sustained long-term expression of endostatin was achieved in lentivirus-transduced orthotopic bladder tumors, and it was associated with decreased vascularization and inhibition of tumor growth. Lentivirus vector-mediated overexpression of endostatin did not affect the intrinsic production of basic fibroblast growth factor and vascular endothelial growth factor.

Conclusions: These findings suggest that lentivirus-mediated gene transfer might represent an effective strategy for expression of angioinhibitory peptides to achieve inhibition of human bladder cancer proliferation and tumor progression.
Histone Deacetylase Inhibitors Have a Profound Antigrowth Activity in Endometrial Cancer Cells
Authors: Takai Noriyuki , Desmond Julian C. , Kumagai Takashi , Gui Dorina , Said Jonathan W. , Whittaker Sadie , Miyakawa Isao , Koeffler H. Phillip
Published: September 2005

Purpose: HDAC inhibitors (HDACIs) have been shown to inhibit cancer cell proliferation, stimulate apoptosis, and induce cell cycle arrest. Our purpose was to investigate the antiproliferative effects of the HDACIs [suberoyl anilide bishydroxamine, valproic acid (VPA), trichostatin A, and sodium butyrate] against six endometrial cancer cell lines.

Experimental Design: Endometrial cancer cells were treated with a variety of HDACIs, and the effect on cell growth, cell cycle, and apoptosis was measured. The ability of VPA to inhibit the growth of endometrial tumors growing in immunodeficient mice was also assessed.

Results: Clonogenic assays showed that all cancer cell lines were sensitive to the growth inhibitory effect of HDACIs. Cell cycle analysis indicated that treatment with HDACIs decreased the proportion of cells in S phase and increased the proportion of cells in the G0-G1 and/or G2-M phases of the cell cycle. Terminal deoxynucleotidyl transferase-mediated nick end labeling assays showed that HDACIs induced apoptosis. This was concomitant with altered expression of genes related to malignant phenotype, including an increase in p21Waf1, p27Kip7, and E-cadherin and a decrease in Bcl-2 and cyclin-D1 and -D2. Chromatin immunoprecipitation analysis revealed a remarkable increase in levels of acetylated histones associated with the p21 promoter after suberoyl anilide bishydroxamine treatment. In nude mice experiments, VPA inhibited significantly human uterine tumor growth without toxic side effects.

Conclusions: These results suggest that HDACIs are effective in inhibiting growth of endometrial cancer cells in vitro and in nude mice, without toxic side effects. The findings raise the possibility that HDACIs may prove particularly effective in treatment of endometrial cancers.
p73 Expression Is Associated with the Cellular Radiosensitivity in Cervical Cancer after Radiotherapy
Authors: Liu Stephanie Si , Leung Rebecca Ching-Yu , Chan Kelvin Yuen-Kwong , Chiu Pui-Man , Cheung Annie Nga-Yin , Tam Kar-Fai , Ng Tong-Yow , Wong Ling-Chui , Ngan Hextan Yuen-Sheung
Published: September 2005

Apoptosis is one of the causes of cell death in cervical cancer following radiotherapy (S. S. Liu et al., Eur. J. Cancer, 37: 1104–1110, 2001). By studying the gene expression profile with cDNA apoptotic array, the p73 gene was found overexpressed in radiosensitive cervical cancers when compared with radioresistant ones. To investigate the role of the p73 gene in relation to clinical assessment of radiosensitivity in cervical cancer based on the findings of residual tumor cells in cervical biopsies after completion of radiotherapy, we studied the protein expression of p73 in 59 cervical cancers after radiotherapy and 68 normal cervices using immunohistochemistry. The expression of p73 was found to be significantly increased in cancer samples and, more importantly, in those samples sensitive to radiotherapy (P < 0.001). The overexpression of p73 actually predicted a better prognosis in cervical cancer patients (P < 0.001). To investigate the possible involvement of p73 downstream genes, the protein expressions of p21 and Bax were studied. The expression of p21, but not Bax, was found to be positively correlated with the expression of p73 (P = 0.001). Furthermore, the epigenetic regulation of p73 expression via DNA methylation was also investigated in 103 cervical cancers and 124 normals. Hypermethylation of p73 gene was observed in 38.8% of cervical cancers, and it was significantly associated with reduced or absent p73 expression (P < 0.001). Reactivation of p73 expression in two cervical cancer cell lines by demethylation treatment supported the role of methylation in the regulation of p73 expression. Our findings suggested that p73 expression was related to the radiosensitivity of cervical cancer cells and may play an important role in the regulation of cellular radiosensitivity.
A Novel Conditionally Replicative Adenovirus Vector Targeting Telomerase-Positive Tumor Cells
Authors: Huang Qian , Zhang Xiuwu , Wang He , Yan Bin , Kirkpatrick John , Dewhrist Mark W. , Li Chuan-Yuan
Published: September 2005

Purpose: To develop a novel conditionally replicative adenovirus vector that targets telomerase-positive cancer cells.

Experimental Design: A telomerase gene-derived promoter was used to control the expression of the E1a gene so that the E1a gene is only expressed in telomerase-positive tumor cells. In addition, a reporter gene was also engineered into the vector so that its infection and replication can be monitored easily.

Results: A novel recombinant adenovirus vector that could selectively replicate in telomerase-positive cancer cells was made successfully. This vector showed active replication in a panel of cancer cells and minimal replication in normal human fibroblast or epithelial cells. The recombinant vector could effectively lyse various cultured tumor cells even at very low multiplicity of infection. The replication efficiency in tumor cells is over 103-fold more than normal fibroblast and epithelial cells. In s.c. tumor models, the newly developed telomerase-selective adenovirus vectors exhibited significantly more virus replication and reporter gene expression.

Conclusions: The telomerase-targeted adenovirus vector has significant potential as an oncolytic virus as well as a tumor-specific therapeutic gene delivery vehicle.
Antisense Oligonucleotide Targeting of Raf-1
Authors: Mullen Peter , McPhillips Fiona , MacLeod Kenneth , Monia Brett , Smyth John F. , Langdon Simon P.
Published: September 2005

Purpose: We sought to identify determinants of growth response to the Raf-1-targeted antisense oligonucleotide (ASO; ISIS 5132) using a large panel of ovarian cancer cell lines.

Experimental Design: First-(ISIS 5132) and second-generation (ISIS 13650) anti-Raf 1 ASOs were compared with control oligonucleotides. Growth was assessed by cell counts; apoptosis was assessed by poly(ADP-ribose) polymerase cleavage; and cell cycle analysis was assessed by flow cytometry. Protein expression was detected by Western blot analysis, and mRNA expression was detected by quantitative reverse transcription-PCR. Raf-1 kinase activity was detected by anti-Raf-1 immunoprecipitation, followed by myelin basic protein phosphorylation.

Results: A panel of 15 ovarian cancer cell lines was used to model a range of growth responses to ASOs targeting Raf-1 mRNA. Growth inhibition varied from 10% to >90% inhibition. Growth inhibition was associated with increased apoptosis and accumulation of cells in the G2-M and S phases of the cell cycle. Growth response was not related to level of Raf-1 protein expression, Raf-1 kinase activity, intracellular ASO uptake, or degree of Raf-1 protein inhibition. However, ASO growth response was associated with a high proportion of Raf-1 mRNA [relative to total (i.e., Raf-1 + A-Raf + B-Raf) Raf mRNA] and significantly higher Raf-1 kinase activity induction following growth factor (transforming growth factor α) stimulation in the cell lines consistent with dependency of these cell lines on Raf-1.

Conclusions: These data indicate that ovarian cancers demonstrate differential sensitivity to ASOs targeted against Raf-1, and target expression levels and degree of utilization of Raf-1 signaling are implicated. Clinically sensitive tumors could feasibly be identified.
Plasma and Cerebrospinal Fluid Pharmacokinetics of Imatinib after Administration to Nonhuman Primates
Authors: Neville Kathleen , Parise Robert A. , Thompson Patrick , Aleksic Alexander , Egorin Merrill J. , Balis Frank M. , McGuffey Leticia , McCully Cynthia , Berg Stacey L. , Blaney Susan M.
Published: September 2005

Purpose: Imatinib mesylate (Gleevec, Glivec, STI571, imatinib) is a potent tyrosine kinase inhibitor approved for the treatment of chronic myelogenous leukemia and gastrointestinal stromal tumors. The role of imatinib in the treatment of malignant gliomas and other solid tumors is being evaluated. We used a nonhuman primate model that is highly predictive of the cerebrospinal fluid penetration of drugs in humans to study the pharmacokinetics of imatinib in plasma and cerebrospinal fluid (CSF) after i.v. and p.o. administration.

Experimental Design: Imatinib, 15 mg/kg i.v. over 30 min (n = 3) or 30 mg/kg p.o. (n = 3), was administered to nonhuman primates. Imatinib was measured in serial samples of plasma and CSF using high-pressure liquid chromatography with UV absorbance or mass spectroscopic detection. Pharmacokinetic parameters were estimated using model-independent methods.

Results: Peak plasma imatinib concentrations ranged from 6.4 to 9.5 μm after i.v. dosing and 0.8 to 2.8 μm after p.o. dosing. The mean ±SD area under the plasma concentration versus time curve was 2480 ±1340 μm·min and 1191 ±146 μm·min after i.v. and p.o. dosing, respectively. The terminal half-life was 529 ±167 min after i.v. dosing and 266 ±88 min after p.o. dosing. After i.v. dosing the steady state volume of distribution was 5.9 ±2.8 liter/kg, and the total body clearance was 12 ±5 ml/min/kg. The mean peak CSF concentration was 0.25 ±0.07 μm after i.v. dosing and 0.07 ±0.04 μm after p.o. dosing. The mean CSF:plasma area under the plasma concentration versus time curve ratio for all of the animals was 5% ±2%.

Conclusions: There is limited penetration of imatinib into the CSF of nonhuman primates after i.v. and p.o. administration.
MUC1 and Nuclear β-Catenin Are Coexpressed at the Invasion Front of Colorectal Carcinomas and Are Both Correlated with Tumor Prognosis
Authors: Baldus Stephan E. , Mönig Stefan P. , Huxel Sandra , Landsberg Stephanie , Hanisch Franz-Georg , Engelmann Katja , Schneider Paul M. , Thiele Jürgen , Hölscher Arnulf H. , Dienes Hans P.
Published: September 2005

Purpose: Overexpression of MUC1 and cytosolic interaction of the mucin with β-catenin are claimed to be involved in colorectal carcinogenesis. In vitro data published recently suggest that MUC1 overexpression results in an increase of steady state levels of nuclear β-catenin. We tried to elucidate the coexpression of both molecules in colorectal cancer to demonstrate possible correlations with clinical, pathological, and prognostic data.

Experimental Design: An immunohistochemical double staining study was performed to characterize the expression and subcellular distribution of MUC1 and β-catenin in a series of 205 patients with colorectal carcinoma. The results were correlated with clinicopathological variables as well as overall survival.

Results: MUC1 was strongly expressed in the tumor center and at the invasion front in ∼50% of the cases. Similar results were obtained with regard to nuclear accumulation of β-catenin at the invasive tumor parts. MUC1 protein expression in the tumor center correlated significantly with a low grade of differentiation, and nuclear β-catenin in the tumor periphery was more frequent in carcinomas of the left colon and rectum. Overexpression of MUC1 and β-catenin, as well as their nuclear coexpression at the invasion front correlated with a worse overall survival in an univariate analysis. However, only pathological tumor-node-metastasis staging and MUC1 at the invasion front revealed as independent prognostic factors.

Conclusions: These results suggest that MUC1 and β-catenin are coexpressed at the invasion front of colorectal carcinomas and that this feature is associated with an accelerated course of disease and worse prognosis.
Evidence for Prostate Cancer-Associated Diagnostic Marker-1
Authors: Ohkia Akira , Hu Youji , Wang Min , Garcia Fernando U. , Stearns Mark E.
Published: September 2005

Purpose: The purpose of this study was to characterize a novel gene/protein associated with prostate cancer, termed prostate cancer-associated diagnostic marker-1 [PCADM-1 (Hu Y, Wang M, Garcia FU, Aoyaki K, Stearns ME. Identification of PCADM-1 as a novel diagnostic marker for prostate cancer, submitted for publication)].

Experimental Design and Results: Immunological studies revealed that rabbit polyclonal antibodies generated against recombinant PCADM-1 specifically recognize the protein in crude protein extracts from a variety of prostate cancer cell lines (i.e., PC-3 ML, LNCaP, DU145, and CPTX-1532) and prostate cancer tissue. Combined immunolabeling and in situ hybridization studies demonstrated that PCADM-1 mRNA was expressed by the luminal epithelial cells of prostate cancer glands and was not expressed by high-grade prostatic intraepithelial neoplasia or HPV-MLC7 cells. Immunolabeling studies of tissue arrays from biopsies of archival material (n = 200 samples) confirmed that PCADM-1 was expressed by the luminal epithelial cells of prostate cancer.

Conclusions: Taken together, the data suggest that PCADM-1 is a specific marker for human prostate cancer.
A Randomized Phase I and Pharmacological Trial of Sequences of 1,3-bis(2-Chloroethyl)-1-Nitrosourea and Temozolomide in Patients with Advanced Solid Neoplasms
Authors: Hammond Lisa A. , Eckardt John R. , Kuhn John G. , Gerson Stanton L. , Johnson Tom , Smith Lon , Drengler Ronald L. , Campbell Elizabeth , Weiss Geoffrey R. , Von Hoff Daniel D. , Rowinsky Eric K.
Published: September 2005

Purpose: O 6-alkylguanine-DNA alkyltransferase (AGAT) is modulated by methylating agents, which, in turn, abrogates nitrosourea resistance in preclinical studies. The feasibility of administering various sequences of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and temozolomide (TEM) in patients with advanced solid neoplasms was evaluated in this Phase I and pharmacological study to assess this premise in the clinical setting. The study also sought to determine the maximum tolerated dose (MTD) levels of BCNU and TEM as a function of Seq, to characterize the pharmacokinetic (PK) behavior of TEM administered both before and after BCNU, assess AGAT fluctuations in peripheral blood mononuclear cells (PBMCs), and seek preliminary evidence of anticancer activity.

Experimental Design: Sixty-three patients were randomized to receive treatment with oral TEM daily on days 1–5 and BCNU administered i.v., either on day 1 before TEM [Sequence (Seq) B→T] or day 5 after TEM (Seq T→B). Treatment was repeated every 6 weeks. Blood sampling for PK studies was performed on both days 1 and 5 of course one. PBMCs were sampled to evaluate major sequence-dependent effects on AGAT levels.

Results: Neutropenia and thrombocytopenia were the principal dose-limiting toxicities of the BCNU/TEM regimen. These effects were more prominent in patients receiving Seq T→B, resulting in a much lower MTD of 80/100 mg/m2/day compared with 150/110 mg/m2/day for Seq B→T. Notable antitumor activity was observed in patients with glioblastoma multiforme, sarcoma, and ovarian carcinoma. No sequence-dependent PK effects were noted to account for sequence-dependent toxicological effects. At the MTD level, AGAT activity in PBMCs decreased 3-fold, on average, and AGAT fluctuations did not appear to be sequence-dependent.

Conclusions: The principal toxicities of the BCNU/TEM regimen were neutropenia and thrombocytopenia, which were consistent and predictable, albeit sequence-dependent. Seq T→B was substantially more myelosuppressive, resulting in disparate MTDs and dose levels recommended for subsequent disease-directed evaluations (150/110 and 80/100 mg/m2/day for Seq B→T and T→B, respectively). Sequence-dependent differences in TEM PK do not account for this clinically relevant magnitude of sequence-dependent toxicity. The characteristics of the myelosuppressive effects of BCNU/TEM, the paucity of severe nonhematological toxicities, and antitumor activity at tolerable doses warrant disease-directed evaluations on this schedule.
Plasminogen Activator Inhibitor-1 as a Potential Marker for the Malignancy of Esophageal Squamous Cell Carcinoma
Authors: Sakakibara Takumi , Hibi Kenji , Kodera Yasuhiro , Ito Katsuki , Akiyama Seiji , Nakao Akimasa
Published: September 2005

Purpose and Experimental Design: To test whether plasminogen activator inhibitor-1 (PAI-1) can serve as a candidate marker for the malignancy of esophageal squamous cell carcinoma (SCC), we performed a quantitative reverse transcription-PCR for PAI-1 gene and evaluated the possible relationship between PAI-1 gene expression levels and clinicopathological findings in esophageal SCC.

Results: Significant increases in PAI-1 scores were observed in metastasis-positive esophageal SCCs (3.08 ± 0.80) compared with metastasis-negative ones (−0.31 ± 0.62; P = 0.0042). PAI-1 expression scores significantly increased with tumor stage (P = 0.05, ANOVA).

Conclusions: These results suggested that PAI-1 might serve as a new parameter for prediction of prognosis in esophageal SCC.
Detection of Prostate Cancer and Predicting Progression
Authors: Tricoli James V. , Schoenfeldt Mason , Conley Barbara A.
Published: September 2005

Carcinoma of the prostate is the second leading cause of male cancer-related death in the United States. Better indicators of prostate cancer presence and progression are needed to avoid unnecessary treatment, predict disease course, and develop more effective therapy. Numerous molecular markers have been described in human serum, urine, seminal fluid, and histological specimens that exhibit varying capacities to detect prostate cancer and predict disease course. However, to date, few of these markers have been adequately validated for clinical use. The purpose of this review is to examine the current status of these markers in prostate cancer and to assess the diagnostic potential for future markers from identified genes and molecules that display loss, mutation, or alteration in expression between tumor and normal prostate tissues. In this review we cite 91 molecular markers that display some level of correlation with prostate cancer presence, disease progression, cancer recurrence, prediction of response to therapy, and/or disease-free survival. We suggest criteria to consider when selecting a marker for further development as a clinical tool and discuss five examples of markers (chromogranin A, glutathione S-transferase π 1, prostate stem cell antigen, prostate-specific membrane antigen, and telomerase reverse transcriptase) that fulfill some of these criteria. Finally, we discuss how to conduct evaluations of candidate prostate cancer markers and some of the issues involved in the validation process.