bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation of Signal Transducer and Activator of Transcription-5 in Prostate Cancer Predicts Early Recurrence
Authors: Li Hongzhen , Zhang Ying , Glass Andrew , Zellweger Tobias , Gehan Edmund , Bubendorf Lukas , Gelmann Edward P. , Nevalainen Marja T.
Published: September 2005

Purpose: We have shown previously that the signal transducer and activator of transcription-5 (Stat5) is a critical survival factor in human prostate cancer cells. In addition, we recently showed that Stat5 is activated at a high level, particularly in high-grade human prostate cancers. Here, we investigated whether activation of Stat5 in prostate cancer was linked to clinical outcome with disease recurrence as end point.

Experimental Design: Immunohistochemistry was used to detect active, nuclear Stat5 in 357 paraffin-embedded prostate cancer specimens on a tissue microarray with clinical follow-up data. Stat5 activation status in prostate cancer specimens was analyzed by univariate and multivariate survival analysis to determine whether activation of Stat5 predicts earlier prostate cancer recurrence. Separate sets of statistical analysis were done for all patients regardless of Gleason grade and for patients with prostate cancer of intermediate Gleason grades (3 and 4).

Results and Conclusions: Stat5 activation in prostate cancer was associated with early disease recurrence (P = 0.0399). Importantly, active Stat5 also predicted shorter progression-free survival in intermediate Gleason grade prostate cancers (P = 0.0409). Stat5 activation remained an independent prognostic marker after adjusting for Gleason grade, pT stage, perineural invasion, or seminal vesicle infiltration in all patients (P = 0.0565) and in Gleason grade 3 or 4 patients (P = 0.0582). The results of this work also confirmed our previous finding of association of Stat5 activation with a high histologic grade of prostate cancer (R = 0.11, P = 0.033). In summary, our study shows that active Stat5 distinguished prostate cancer patients whose disease is likely to progress earlier; therefore, active Stat5 may be a useful marker for selection of more individualized treatment. The results of this study need to be validated in a large prospective cohort.
Promoter Hypermethylation Is the Predominant Mechanism in hMLH1 and hMSH2 Deregulation and Is a Poor Prognostic Factor in Nonsmoking Lung Cancer
Authors: Hsu Han-Shui , Wen Chiao-Kai , Tang Yen-An , Lin Ruo-Kai , Li Wing-Yin , Hsu Wen-Hu , Wang Yi-Ching
Published: September 2005

Purpose and Experimental Design: The etiologic association and prognostic significance of mismatch repair gene/protein alterations have never been examined in nonsmoking lung cancer. Therefore, we investigated protein expression and promoter hypermethylation of hMLH1 and hMSH2 genes in the tumor specimens from 105 nonsmoking female non–small cell lung cancer (NSCLC) patients. Immunohistochemistry and restriction enzyme–based multiplex PCR were used to examine the protein expression and promoter hypermethylation, respectively. The occurrence of gene/protein alteration for each gene was compared with the patients' clinicopathologic variables as well as the overall survival and cancer-specific survival rates.

Results: Protein expression alteration and promoter hypermethylation were observed in 66% to 67% and 30% to 34% of tumor specimens for hMLH1 and hMSH2 genes, respectively. Loss of hMLH1 and hMSH2 protein expression was significantly associated with their promoter hypermethylation (P < 0.0001 and P = 0.049). The overall survival and cancer-specific survival rates were significantly lower in patients with promoter hypermethylation of hMSH2 gene than in those without hypermethylation (P = 0.038 and P = 0.004). The poor prognosis was still especially significant in adenocarcinoma (P = 0.035 and P = 0.061) and early-stage NSCLC patients (P = 0.067 and P = 0.041).

Conclusion: Our data suggest that hMLH1 is the major altered mismatch repair gene involved in nonsmoking NSCLC tumorigenesis and that promoter methylation is the predominant mechanism in hMLH1 and hMSH2 deregulation. In addition, promoter methylation of the hMSH2 gene may be a potential prognostic factor in nonsmoking female lung cancer.
Roxithromycin Inhibits Constitutive Activation of Nuclear Factor κB by Diminishing Oxidative Stress in a Rat Model of Hepatocellular Carcinoma
Authors: Ueno Shinichi , Aoki Dai , Kubo Fumitake , Hiwatashi Kiyokazu , Matsushita Kenji , Oyama Tohru , Maruyama Ikuro , Aikou Takashi
Published: September 2005

Purpose: Recently, 14-member macrolide antibiotics such as clarithromycin and roxithromycin have been shown to have anticancer and antiangiogenic effects. We investigated the suppressive effect of roxithromycin on accelerated hepatocellular carcinoma growth in a rat hepatocarcinogenetic model and compared results with effects from TNP-470.

Experimental Design: Tumor was induced by oral diethylnitrosamine administration for 17 weeks. Normal saline, TNP-470 (50 mg/kg), or roxithromycin (40 or 100 mg/kg) was administered i.p. thrice per week from week 10 to 17.

Results: Carcinomatous tissue growing outside dysplastic nodules and a marked expression of placental glutathione S-transferase were detected in rats with induced carcinogenesis. Tumor growth was accompanied by augmented expression of inducible nitric oxide synthase, activation of nuclear factor κB, and increased lipid peroxidation level. All these effects were absent in animals that received roxithromycin or TNP-470. The inhibitory effect of roxithromycin was dose dependent and no clear differences were noted between groups given roxithromycin 100 mg/kg and TNP-470 50 mg/kg.

Conclusions: Our results indicate that roxithromycin inhibits oxidative stress, nitric oxide production, and nuclear factor κB activation induced by experimental hepatocarcinogenesis. The data provide additional evidence for the potential use of roxithromycin in treatment of hepatocellular carcinoma prevention.
Possible Involvement of RUNX3 Silencing in the Peritoneal Metastases of Gastric Cancers
Authors: Sakakura Chouhei , Hasegawa Kazunori , Miyagawa Koji , Nakashima Susumu , Yoshikawa Tetsuji , Kin Shuichi , Nakase Yuenn , Yazumi Shujiro , Yamagishi Hisakazu , Okanoue Takeshi , Chiba Tsutomu , Hagiwara Akeo
Published: September 2005

Purpose: Our previous results suggested that a lack of RUNX3 function contributed to human gastric carcinogenesis, but the role of RUNX3 in progression and metastasis remains unclear. We examined RUNX3 expression in clinical samples of peritoneal metastases in gastric cancers. Changes in metastatic potential were assessed in animal experiments using stable RUNX3 transfectants of gastric cancer cells. Finally, global expression changes were analyzed using a cDNA microarray.

Experimental Design and Results: Significant down-regulation of RUNX3 through methylation on the promoter region was observed in primary tumors (75%) as well as in all clinical peritoneal metastases of gastric cancers (100%) compared with normal gastric mucosa. Stable transfection of RUNX3 inhibited cell proliferation slightly, and modest transforming growth factor-β (TGF-β)–induced antiproliferative and apoptotic effects were observed. Interestingly, it strongly inhibited peritoneal metastases of gastric cancers in animal model (P < 0.01). Furthermore, we did globally analyzed expression profiles of ∼21,000 genes in parent cells and stable transfectant of RUNX3 using a cDNA microarray. Microarray analysis identified ∼28 candidate genes under the possible downstream control of RUNX3, some of these genes were considered to be possibly involved in peritoneal metastases, which were related to signal transduction (vav3, TOLL-like receptor, MAPKK, MET, S1 00A1 1, and cathepsin E), apoptosis (caspase 9), immune responses (CD55 and TLR1O), and cell adhesion (sialyltransferase 1 and galectin 4). Some of the genes are involved in the TGF-β signaling pathway.

Conclusion: These results indicate that silencing of RUNX3 affects expression of important genes involved in aspects of metastasis including cell adhesion, proliferation, apoptosis, and promoting the peritoneal metastasis of gastric cancer. Identification of such genes could suggest new therapeutic modalities and therapeutic targets.
Phase I and Pharmacologic Study of Infusional Topotecan and Carboplatin in Relapsed and Refractory Acute Leukemia
Authors: Kaufmann Scott H. , Karp Judith E. , Letendre Louis , Kottke Timothy J. , Safgren Stephanie , Greer Jackie , Gojo Ivana , Atherton Pamela , Svingen Phyllis A. , Loegering David A. , Litzow Mark R. , Sloan Jeff A. , Reid Joel M. , Ames Matthew M. , Adjei Alex A. , Erlichman Charles
Published: September 2005

Purpose: To assess the maximum tolerated dose, toxicities, pharmacokinetics, and antileukemic activity of topotecan and carboplatin in adults with recurrent or refractory acute leukemias.

Experimental Design: Patients received topotecan and carboplatin by 5-day continuous infusion at nine dose levels. Patients achieving a complete remission received up to two additional courses for consolidation. Plasma topotecan and ultrafilterable platinum were assayed on days 1 to 5. In addition, pretreatment levels of various polypeptides in leukemic cells were examined by immunoblotting to assess possible correlations with response.

Results: Fifty-one patients received a total of 69 courses of therapy. Dose-limiting toxicity consisted of grade 4/5 typhlitis and grade 3/4 mucositis after one course of therapy or grade 4 neutropenia and thrombocytopenia lasting >50 days when a second course was administered on day 21. Among 45 evaluable patients, there were 7 complete remissions, 2 partial remissions, 1 incomplete complete remission, and 1 reversion to chronic-phase chronic myelogenous leukemia. Topotecan steady-state plasma concentrations increased with dose. No accumulation of topotecan or ultrafilterable platinum occurred between days 1 and 5 of therapy. Leukemic cell levels of topoisomerase I, checkpoint kinase 1, checkpoint kinase 2, and Mcl-1 correlated with proliferating cell nuclear antigen but not with response. In contrast, low Bcl-2 expression correlated with response (P = 0.014, Mann-Whitney U test).

Conclusions: The maximum tolerated dose was 1.6 mg/m2/d topotecan plus 150 mg/m2/d carboplatin. The complete remission rate in a heavily pretreated population was 16% (33% at the highest three dose levels). Responses seem to correlate with low pretreatment blast cell Bcl-2 expression.
Sequential Platinum-Based Chemotherapy-Thoracic Radiotherapy in Early Stage Non–Small Cell Lung Cancer
Authors: Thatcher Nick , Faivre-Finn Corinne , Blackhall Fiona , Anderson Heather , Lorigan Paul
Published: September 2005

Combined chemoradiotherapy (CTRT) is intended to reduce the risk of local regional recurrence and distant relapse in patients with early stage non–small cell lung cancer (NSCLC). Sequential CTRT allows full doses of each modality while avoiding the additive toxicity that occurs with concurrent therapy. This review will encompass studies in the three main settings where sequential CTRT has been applied in early stage NSCLC: as adjuvant therapy after resection, as neoadjuvant chemotherapy and surgery followed by radiotherapy, and in unresectable stage III disease.
Future of Early Detection of Lung Cancer: The Role of Mouse Models
Authors: Shaw Alice T. , Kirsch David G. , Jacks Tyler
Published: September 2005

Early detection represents one of the most promising approaches to improving lung cancer survival. To date, no screening strategies have been shown to decrease mortality from the disease. Furthermore, no reliable circulating biomarkers of lung cancer have been identified that allow early diagnosis. With the advent of gene targeting technology, new genetically engineered mouse models of lung cancer closely recapitulate the pathobiology of human disease. These mouse models have enabled novel approaches to early detection, including the identification of cancer-associated serum markers using proteomic technologies and the development of new molecular imaging tools. The application of innovative technologies to accurate mouse models promises to accelerate the discovery of new molecular targets and imaging biomarkers for the early detection of lung cancer.
Adjuvant Chemotherapy for Non–Small Cell Lung Cancer: Contribution of the International Adjuvant Lung Trial
Authors: Dunant Ariane , Pignon Jean-Pierre ,   Thierry Le Chevalier on behalf of the International Adjuvant Lung Cancer Trial Collaborative Group
Published: September 2005

The recently reported International Adjuvant Lung Cancer Trial (IALT) was designed to assess the potential benefit of three to four cycles of adjuvant cisplatin-based chemotherapy after complete resection of non–small cell lung cancer (NSCLC). Each center predetermined the cisplatin dose (total 300-400 mg/m2), the combined drug (etoposide or a Vinca alkaloid), and the radiotherapy policy. From 1995 to 2000, 1,867 patients were randomized in 148 centers from 33 countries. On September 1, 2002, median follow-up was 56 months and >98% of patients had an updated follow-up. Overall survival was significantly different between the two arms: 5-year survival rate was 44.5% in the chemotherapy arm versus 40.4% in the control arm [relative risk = 0.86 (0.76-0.98), P < 0.03]. Disease-free survival, incidence of local recurrence, and incidence of distant metastases (but not brain metastases) were likewise significantly different, with an advantage for the chemotherapy arm. We concluded that adjuvant cisplatin-based chemotherapy in resected NSCLC should become part of the standard management of operable NSCLC. Two other recently reported randomized prospective studies also showed a significant benefit for postoperative platin-based doublets in stage IB and II NSCLC and confirmed the role of adjuvant chemotherapy as part of the treatment of these patients. The Lung Adjuvant Cisplatin Evaluation program, a pooled analysis of all recent platin-based adjuvant trials, and the IALT-Bio study, which will investigate over 30 markers in the IALT patients' specimens, should allow us to better define the populations more likely to benefit from postoperative chemotherapy.
The ALPI Trial: The Italian/European Experience with Adjuvant Chemotherapy in Resectable Non–Small Lung Cancer
Authors: Scagliotti Giorgio V. ,   on behalf of the Adjuvant Lung Cancer Project Italy/European Organization for Research and Treatment of Cancer Lung Cancer Cooperative Group
Published: September 2005

Postoperative treatments for lung cancer have been evaluated for more than two decades, but in the majority of the studies no significant and clinically meaningful effect on survival has been shown. In 1995, a meta-analysis of eight cisplatin-based adjuvant chemotherapy trials in 1,394 patients with non–small cell lung cancer showed a 13% reduction in the risk of death (P = 0.08). The nonstatistically significant benefit reported in the meta-analysis prompted the planning of several randomized studies of platinum-based chemotherapy. Three studies addressed the issue of adjuvant chemotherapy in all the resected stages of non–small cell lung cancer (I-IIIA): the Italian/European study Adjuvant Lung Cancer Project Italy, the International Adjuvant Lung Cancer study, and the British Big Lung Trial. In contrast to the International Adjuvant Lung Cancer, the Adjuvant Lung Cancer Project Italy and the underpowered British Big Lung Trial failed to prospectively confirm a significant role of adjuvant chemotherapy in completely resected non–small cell lung cancer. In this article, we will discuss the findings of the Adjuvant Lung Cancer Project Italy study in the context of the International Adjuvant Lung Cancer and British Big Lung Trial.
Gene Expression Profiling of Renal Cell Carcinoma
Authors: Tan Min-Han , Rogers Craig G. , Cooper Jeffrey T. , Ditlev Jonathon A. , Maatman Thomas J. , Yang Ximing , Furge Kyle A. , Teh Bin Tean
Published: September 2005

Renal cell carcinoma (RCC) is a histologically diverse disease, with variable and often unpredictable clinical behavior. The prognosis worsens dramatically with the onset of clinical metastasis, and current regimens of systemic therapy yield only modest benefits for metastatic RCC. Gene expression profiling is a promising technique for refining the diagnosis and staging of RCC, as well as for highlighting potential therapeutic targets. We review the recent advances in expression profiling of RCC and discuss the clinical and biological insights obtained from these studies.
Review of Radiofrequency Ablation for Renal Cell Carcinoma
Authors: Hines-Peralta Andrew , Goldberg S. Nahum
Published: September 2005

This review will discuss how minimally invasive, image-guided radiofrequency (RF) tumor ablation [i.e., coagulating tumor using short-duration heating (<15 minutes) by directly applying temperatures >50°C via needle electrodes] is being incorporated as a clinical tool for the treatment of renal cell carcinoma. RF ablation has been used to treat focal liver tumors. Potential benefits of this thermal therapy include reduced morbidity and mortality compared with standard surgical resection and the ability to treat nonsurgical patients. More recently, this technique has been introduced to treat focal renal tumors, particularly incidental lesions smaller than 3 cm in elderly patients and those with comorbid conditions. Other uses have included treatment in patients with von Hippel-Lindau syndrome and other diseases that predispose patients to multiple renal carcinomas, where renal parenchymal preservation is desired. Techniques, complications, and results will be discussed. Additionally, strategies that we are currently studying to improve RF outcomes and enable the potential treatment of larger tumors will be addressed. Most notably, recent data on increased coagulation achieved by combining RF ablation with antivascular/antiangiogenic therapies, such as arsenic trioxide, that reduce blood flow and promote heat retention are provided.
Bevacizumab for Patients with Metastatic Renal Cancer
Authors: Yang James C.
Published: September 2005

Most clear cell renal cell cancer (RCC) is caused by biallelic loss of the von Hippel-Lindau gene. One consequence of this loss is up-regulation of vascular endothelial growth factor via a pathway involving accumulation of hypoxia inducible factor. Vascular endothelial growth factor, a potent angiogenic factor, is secreted by many human cancers, but clear cell RCC as a group produces particularly high levels and has a highly vascular histologic appearance. In a randomized, placebo-controlled, double-blind trial, we tested the use of a neutralizing antibody to vascular endothelial growth factor, bevacizumab, in patients with metastatic RCC. At 3 or 10 mg/kg every 2 weeks, toxic effects were minimal, with hypertension and proteinuria the most substantial events. There were four partial responses (10% response rate) and a highly substantial prolongation of time to tumor progression in patients who received the higher dose of bevacizumab. With a crossover design and very sensitive criteria for disease progression, no difference in survival was shown. Four patients have been undergoing long-term bevacizumab therapy without tumor progression for 3 to 5 years. Three have substantial proteinuria but retain normal renal function. A small pilot trial combining bevacizumab and thalidomide showed no unexpected toxic effects. Future trials should consider combination therapies and strategies in which patients are treated through initial disease progression with antiangiogenic agents such as bevacizumab.
Molecular Markers for Early Detection of Renal Carcinoma
Authors: Skates Steven , Iliopoulos Othon
Published: September 2005

Molecular markers for renal cell carcinoma could guide early detection of localized disease in defined populations at high risk for the disease or early disease recurrence after nephrectomy for renal cell carcinoma. In addition, sensitive and specific markers may provide surrogate end points for clinical trials of treatment and/or disease prevention. Powerful techniques of genomic and proteomic analysis of human renal carcinoma cell lines, tumor samples, and biological fluids, such as plasma and urine, obtained from patients with renal cell carcinoma, are likely to identify candidate markers. Careful selection among early candidate markers and further testing in independent patient populations are required for marker validation.
KIT (CD117)-Positive Breast Cancers Are Infrequent and Lack KIT Gene Mutations
Authors: Simon Ronald , Panussis Soti , Maurer Robert , Spichtin Hanspeter , Glatz Kathrin , Tapia Coya , Mirlacher Martina , Rufle Alex , Torhorst Joachim , Sauter Guido
Published: September 2005

Purpose: KIT (CD117) is a transmembrane tyrosine kinase representing a target for STI571 (Glivec) therapy. Some KIT-overexpressing solid tumors have responded favorably to STI571, potentially because of the presence of KIT-activating mutations.

Experimental Design: To investigate the epidemiology of KIT overexpression and mutations, we investigated a series of 1654 breast cancers. All tumors were analyzed by immunohistochemistry in a tissue microarray format.

Results: KIT expression was always present in normal breast epithelium. However, cancer analysis revealed the only 43 of 1654 (2.6%) tumors were KIT-positive. KIT expression was more frequent in medullary cancer (9 of 47 positive; 19.1%) than in any other histological tumor subtype (P < 0.001). KIT expression was significantly associated with high tumor grade (P < 0.0001) but unrelated to pT and pN categories or patient survival. Mutation analysis of exons 2, 8, 9, 11, 13, and 17 was negative in 10 KIT-positive tumors.

Conclusions: Overall, our data show that a high level of KIT expression occurs infrequently in breast cancer. KIT-positive breast cancers may not reflect “KIT up-regulation” because KIT is also expressed in normal breast epithelium. The lack of KIT mutations also argues against the therapeutic efficacy of STI571 in breast cancer.
Altered N-myc Downstream-Regulated Gene 1 Protein Expression in African-American Compared with Caucasian Prostate Cancer Patients
Authors: Caruso Robert P. , Levinson Benjamin , Melamed Jonathan , Wieczorek Rosemary , Taneja Samir , Polsky David , Chang Caroline , Zeleniuch-Jacquotte Anne , Salnikow Konstantin , Yee Herman , Costa Max , Osman Iman
Published: September 2005

Purpose: The protein encoded by N-myc downstream-regulated gene 1 (NDRG1) is a recently discovered protein whose transcription is induced by androgens and hypoxia. We hypothesized that NDRG1 expression patterns might reveal a biological basis for the disparity of clinical outcome of prostate cancer patients with different ethnic backgrounds.

Experimental Design: Patients who underwent radical prostatectomy between 1990 and 2000 at Veterans Administration Medical Center of New York were examined. We studied 223 cases, including 157 African Americans and 66 Caucasians (T2, n = 144; ≥T3, n = 79; Gleason <7, n = 122; ≥7, n = 101). Three patterns of NDRG1 expression were identified in prostate cancer: (a) intense, predominately membranous staining similar to benign prostatic epithelium; (b) intense, nucleocytoplasmic localization; and (c) low or undetectable expression. We then examined the correlations between patients’ clinicopathological parameters and different NDRG1 expression patterns.

Results: In this study of patients with equal access to care, African-American ethnic origin was an independent predictor of prostate-specific antigen recurrence (P < 0.05). We also observed a significant correlation between different patterns of NDRG1 expression and ethnic origin. Pattern 2 was less frequent in African Americans (21% versus 38%), whereas the reverse was observed for pattern 3 (60% in African Americans versus 44% in Caucasians; P = 0.03). This association remained significant after controlling for both grade and stage simultaneously (P = 0.02).

Conclusions: Our data suggest that different NDRG1 expression patterns reflect differences in the response of prostatic epithelium to hypoxia and androgens in African-American compared with Caucasian patients. Further studies are needed to determine the contribution of NDRG1 to the disparity in clinical outcome observed between the two groups.
Genomic Organization, Incidence, and Localization of the SPAN-X Family of Cancer-Testis Antigens in Melanoma Tumors and Cell Lines
Authors: Westbrook V. Anne , Schoppee Pamela D. , Diekman Alan B. , Klotz Kenneth L. , Allietta Margaretta , Hogan Kevin T. , Slingluff Craig L. , Patterson James W. , Frierson Henry F. , Irvin William P. , Flickinger Charles J. , Coppola Michael A. , Herr John C.
Published: September 2005

Purpose: Members of the SPAN-X (sperm protein associated with the nucleus mapped to the X chromosome) family of cancer-testis antigens are promising targets for tumor immunotherapy because they are normally expressed exclusively during spermiogenesis on the adluminal side of the blood-testis barrier, an immune privileged compartment.

Experimental Design and Results: This study analyzed the human SPANX genomic organization, as well as SPAN-X mRNA and protein expression in somatic and cancer cells. The SPANX family consists of five genes, one of which is duplicated, all located in a gene cluster at Xq27.1. From the centromere, the arrangement of the five SPANX genes mapped on one contiguous sequence is SPANXB, -C, -A1, -A2, and -D. Reverse transcription-PCR analyses demonstrated expression of SPAN-X mRNA in melanoma and ovarian cell lines, and virtual Northern analysis established SPANX gene expression in numerous cancer cell lines. Immunoblot analysis using polyclonal antisera raised against recombinant SPAN-X confirmed the translation of SPAN-X proteins in melanoma and ovarian tumor cell lines. The immunoreactive proteins migrated between Mr 15,000 and Mr 20,000 similar to those observed in spermatozoa. Immunoperoxidase labeling of melanoma cells and tissue sections demonstrated SPAN-X protein localization in the nucleus, cytoplasm, or both. Ultrastructurally, in melanoma cells with nuclear SPAN-X, the protein was associated with the nuclear envelope, a localization similar to that observed in human spermatids and spermatozoa. Significantly, the incidence of SPAN-X-positive immunostaining was greatest in the more aggressive skin tumors, particularly in distant, nonlymphatic metastatic melanomas.

Conclusions: The data herein suggest that the SPAN-X protein may be a useful target in cancer immunotherapy.
Src Kinase and Mitogen-Activated Protein Kinases in the Progression from Normal to Malignant Endometrium
Authors: Desouki Mohamed Mokhtar , Rowan Brian G.
Published: September 2005

Purpose: The purpose of this research was to determine whether a correlation exists between the levels of activated mitogen-activated protein kinase (MAPK) and Src kinases and the progression from normal to malignant endometrium.

Experimental Design: We measured total and phosphorylated levels for extracellular signal-regulated kinase 1/2, p38, stress-activated protein kinase/c-Jun NH2-terminal kinase, and Src kinases from 33 frozen endometrial adenocarcinomas and 38 benign endometrial specimens by quantitation of signals from Western blots using antibodies against these kinases.

Results: Elevated phospho-extracellular signal-regulated kinase 1/2 (150 ± 40 versus 46 ± 7; P = 0.03), phospho-Src (28 ± 5 versus 4 ± 1), and phospho-p38 (131 ± 16 versus 27 ± 7; P < 0.001) was detected in benign versus malignant endometrium when the Western blot signal of activated kinase was normalized to total kinase levels and β actin. A modest increase in active c-Jun NH2-terminal kinase was detected in carcinoma versus benign specimens (51 ± 13 versus 43 ± 10; P = 0.8). Expression of total kinases (normalized to β-actin) was higher in carcinoma versus benign specimens, respectively (extracellular signal-regulated kinase 1/2, 9 ± 2 versus 0.7 ± 0.1; Src, 7 ± 2 versus 0.4 ± 0.1; stress-activated protein kinase c-Jun NH2-terminal kinase, 2 ± 0.4 versus 0.2 ± 0.02; P < 0.001; and p38, 1 ± 0.2 versus 0.4 ± 0.1; P < 0.01). Immunohistochemistry for active and total Src kinases and MAPKs detected positive staining in epithelial and stroma cells.

Conclusions: These data demonstrated that, in contrast with breast cancer, the progression from normal to malignant endometrium is not associated with activation of MAPK and Src kinases. Elevation of these active kinases in benign endometrium may contribute to endometrial resistance to the antiestrogen action of tamoxifen.
Introduction of Human Telomerase Reverse Transcriptase to Normal Human Fibroblasts Enhances DNA Repair Capacity
Authors: Shin Ki-Hyuk , Kang Mo K. , Dicterow Erica , Kameta Ayako , Baluda Marcel A. , Park No-Hee
Published: September 2005

Purpose: From numerous reports on proteins involved in DNA repair and telomere maintenance that physically associate with human telomerase reverse transcriptase (hTERT), we inferred that hTERT/telomerase might play a role in DNA repair. We investigated this possibility in normal human oral fibroblasts (NHOF) with and without ectopic expression of hTERT/telomerase.

Experimental Design: To study the effect of hTERT/telomerase on DNA repair, we examined the mutation frequency rate, host cell reactivation rate, nucleotide excision repair capacity, and DNA end-joining activity of NHOF and NHOF capable of expressing hTERT/telomerase (NHOF-T). NHOF-T was obtained by transfecting NHOF with hTERT plasmid.

Results: Compared with parental NHOF and NHOF transfected with empty vector (NHOF-EV), we found that (a) the N-methyl-N′-nitro-N-nitrosoguanidine-induced mutation frequency of an exogenous shuttle vector was reduced in NHOF-T, (b) the host cell reactivation rate of N-methyl-N′-nitro-N-nitrosoguanidine-damaged plasmids was significantly faster in NHOF-T; (c) the nucleotide excision repair of UV-damaged DNA in NHOF-T was faster, and (d) the DNA end-joining capacity in NHOF-T was enhanced. We also found that the above enhanced DNA repair activities in NHOF-T disappeared when the cells lost the capacity to express hTERT/telomerase.

Conclusions: These results indicated that hTERT/telomerase enhances DNA repair activities in NHOF. We hypothesize that hTERT/telomerase accelerates DNA repair by recruiting DNA repair proteins to the damaged DNA sites.
Enhanced Protein Profiling Arrays with ELISA-Based Amplification for High-Throughput Molecular Changes of Tumor Patients’ Plasma
Authors: Huang Ruochun , Lin Ying , Shi Qian , Flowers Lisa , Ramachandran Sumathi , Horowitz Ira R. , Parthasarathy Sampath , Huang Ruo-Pan
Published: September 2005

Purpose: The purpose of this study is to develop a high-throughput approach to detect protein expression from hundreds and thousands of samples and to apply this technology to profile circulating angiogenic factor protein levels in patients with gynecological tumors.

Experimental Design: Analytes containing a mixture of protein are immobilized onto antibody-coated surface of support in array format. The presence of protein in analytes is detected with biotin-labeled antibody coupled with an enhanced chemiluminescence or fluorescence detection system. The exact amount of protein can be quantitatively measured. The expression levels of five angiogenic factors (angiogenin, interleukin 8, vascular endothelial growth factor, platelet-derived growth factor, and epidermal growth factor) from 157 samples were quantitatively measured using this novel protein array technology and were statistically analyzed. The expression patterns of angiogenic factors were analyzed using two-way hierarchical cluster analysis approach.

Results: A novel protein array technology, which can simultaneously and quantitatively measure few protein levels from hundreds and thousands of samples was developed. Only minute amounts of sample are required for the assay. This approach also features high sensitivity and specificity. Using this novel protein array approach, we analyzed the plasma expression levels of five angiogenic factors in 137 patients diagnosed with a tumor and 20 controls. Statistical analysis reveals different expression levels of angiogenic factors between patients and controls. Cluster analysis suggests a possible classification of normal subjects from patients.

Conclusions: Enhanced protein profiling arrays provide a high-throughput and sensitive system to detect one or few protein from hundreds and thousands of samples. Such an approach should have broad application in biomedical discovery.
Mitochondrial DNA Mutation at the D310 (Displacement Loop) Mononucleotide Sequence in the Pathogenesis of Gallbladder Carcinoma
Authors: Tang Moying , Baez Sergio , Pruyas Martha , Diaz Alfonso , Calvo Alfonso , Riquelme Erick , Wistuba Ignacio I.
Published: September 2005

Purpose: Mutations in the mitochondrial DNA (mtDNA) have been observed frequently in human neoplasia, in both coding and noncoding regions. A mononucleotide repeat (poly-C) between 303 and 315 nucleotides (D310) within the regulatory displacement loop has been identified recently as a frequent hot spot of deletion/insertion mutations in tumors. We investigated the frequency and pattern of D310 abnormalities in the pathogenesis of gallbladder carcinoma (GBC).

Experimental Design: DNA extracted from neoplastic and nonneoplastic archival gallbladder tissue including 123 tumors, 53 dysplastic areas, and 90 histologically normal epithelia adjacent to GBC, chronic cholecystitis, and 15 normal gallbladders were examined by PCR-based assay for D310 mutations, followed by sequencing in a subset of cases.

Results: D310 mutation was a relatively frequent (47 of 123; 38%) abnormality in GBC. A very high frequency of mutations were detected in dysplastic (8 of 14; 57%) and normal-appearing gallbladder epithelia (10 of 22; 46%) accompanying GBC, showing a clonal relationship compared with the corresponding tumors. D310 mutations were also detected in dysplastic (8 of 39; 21%) and normal (17 of 68; 25%) epithelia obtained from chronic cholecystitis. A single case of 15 normal gallbladders showed a D310 abnormality. Overall, deletions (67 of 91; 74%) at D310 were more frequent than insertions.

Conclusions: D310 mutation at the mtDNA displacement loop is a relatively frequent and early event in the sequential pathogenesis of GBC, being detected in normal-appearing epithelium from chronic cholecystitis. Our findings suggest that mtDNA mutations should be additionally investigated in GBC pathogenesis, and D310 mononucleotide abnormalities could be included in a panel of molecular biomarkers for GBC early detection strategy.
Membranous Expression of Secreted Frizzled-Related Protein 4 Predicts for Good Prognosis in Localized Prostate Cancer and Inhibits PC3 Cellular Proliferation
in Vitro
Authors: Horvath Lisa G. , Henshall Susan M. , Kench James G. , Saunders Darren N. , Lee C.-Soon , Golovsky David , Brenner Phillip C. , O’Neill Gordon F. , Kooner Raji , Stricker Phillip D. , Grygiel John J. , Sutherland Robert L.
Published: September 2005

Purpose: Activation of the Wnt-signaling pathway is implicated in aberrant cellular proliferation in a variety of cancers. Secreted frizzled-related protein 4 (sFRP4) is a secreted protein with putative inhibitory activity of the Wnt-signaling cascade through binding and sequestering Wnt ligands. Because sFRP4 mRNA is overexpressed in prostate cancers (PCs), the aim of this study was to define the pattern of sFRP4 protein expression in normal and malignant human prostate tissue and to determine whether changes in expression were associated with disease progression and prognosis, as well as to define the phenotype of sFRP4-overexpression in an in vitro model of PC.

Experimental Design: Polyclonal antibodies were raised against a COOH-terminal peptide of sFRP4, characterized and used to assess sFRP4 protein expression in benign prostate tissue and 229 patients with clinically localized PC (median follow-up 77 months, range 1–156). In vitro studies of the function of sFRP4 overexpression were performed using PC3 cells transfected with sFRP4.

Results: Benign and malignant prostate tissue demonstrated cytoplasmic sFRP4 immunoreactivity, but there was a decrease in the expression of membranous sFRP4 in PCs compared with the hyperplastic lesions (P < 0.0001). Kaplan-Meier analysis revealed that patients whose PC expressed membranous sFRP4 in >20% of cells had improved relapse-free survival compared with those with ≤20% membranous expression (P = 0.002). Moreover, membranous sFRP4 expression (P = 0.04) was an independent predictor of relapse when modeled with Gleason score (P = 0.006), pathological stage (P = 0.002), and pre-operative prostate-specific antigen levels (P = 0.004). In addition, in vitro studies demonstrated a decrease in the proliferation rate of PC3 cells transfected with sFRP4 when compared with the control PC3-empty vector cells (P < 0.0001). Decreased levels of phosphorylated glycogen synthase kinase 3β in PC3-sFRP4 cells suggested that this phenotype is mediated by the “Wnt/β-catenin” pathway.

Conclusions: These data suggest that sFRP4 expression may be prognostic for localized PC, potentially as a consequence of an inhibitory effect on PC cell proliferation.
Degree of CD25 Expression in T-Cell Lymphoma Is Dependent on Tissue Site
Authors: Jones Dan , Ibrahim Sherif , Patel Kaushali , Luthra Rajyalakshmi , Duvic Madeleine , Medeiros L. Jeffrey
Published: September 2005

Purpose: Using concurrent tumor samples from different anatomical sites, we compared expression of the therapeutic targets CD25 and CD30 in T-cell lymphoma (TCL).

Experimental Design: We examined levels of CD25 and CD30 by flow cytometry in tumor cells from peripheral blood and lymph node in 13 cutaneous TCL patients and by immunohistochemistry in concurrent lymph node and skin biopsy specimens in 17 additional TCL cases, mostly mycosis fungoides. Tumor cell expression was correlated with patterns of expression in nonneoplastic lymphocytes in 14 reactive lymph node and 10 skin samples showing chronic dermatitis. Expression of CD25 and CD30 in all biopsy samples was compared with that of cutaneous lymphocyte antigen (CLA), a mediator of skin homing.

Results: By flow cytometry, we noted significantly decreased expression of CD25 in lymph node compared with peripheral blood in 8 of 13 TCLs, with no changes in CD30 levels in 4 cases studied. Using immunohistochemistry, CD25 was strongly expressed in epidermotropic tumor cells in 13 of 17 (76%) TCL skin specimens but was decreased in the corresponding lymph node in 12 of these cases. CD30 was expressed at roughly equal intensity in tumor cells from both sites, except in 1 case. CLA showed a similar pattern to CD25, being expressed by tumor cells in 16 of 17 (94%) skin specimens, but was largely absent in tumor cells in the corresponding lymph node in 12 of these patients. In T cells from reactive lymph node biopsy specimens, CD25 was highly expressed only in dermatopathic lymphadenitis associated with transient skin rashes.

Conclusions: We demonstrate in vivo that decreased levels of CD25 expression occur in TCL when it involves lymph node, similar to what is seen with CLA. This demonstrable variation related to anatomical localization has implications for the measurement of surface expression of CD25 and for understanding the response of patients with cutaneous TCL to interleukin 2 receptor-targeted immunotherapy.
CD26 Expression Correlates with a Reduced Sensitivity to 2′-Deoxycoformycin-Induced Growth Inhibition and Apoptosis in T-Cell Leukemia/Lymphomas
Authors: Aldinucci Donatella , Poletto Dalisa , Lorenzon Debora , Nanni Paola , Degan Massimo , Olivo Karin , Rapanà Barbara , Pinto Antonio , Gattei Valter
Published: September 2005

Purpose and Experimental Design: dCF (2′-deoxycoformycin) is a potent inhibitor of ADA (adenosine deaminase), an enzyme regulating intra- and extracellular concentrations of purine metabolites. ADA exists as cytosolic and extracellular forms, the latter colocalized on the cell surface with CD26. Once the surface expression of CD26 and ADA in a panel of cell lines and primary samples of T-cell leukemia/lymphoma was defined, we correlated this expression with the antiproliferative and apoptotic effect of dCF.

Results: Surface expression of CD26 inversely correlated with the capability of dCF to inhibit cell growth and induce apoptosis both in T-cell lines and primary samples of T-cell malignancies. This conclusion was sustained by a decreased sensitivity to dCF-mediated proapoptotic and/or antiproliferative in vitro effects of: (a) leukemia/lymphoma T-cell lines expressing surface CD26/ADA complex; (b) primary CD26+ T cell malignancies; and (c) normal T cells (CD26+) as compared with tumor T cells (CD26−) in unpurified samples from three cases of T-cell receptor γδ+ T-cell malignancies characterized by a mixture of normal and neoplastic cells. This latter point was confirmed in vivo, in a patient affected by CD26− T-cell receptor γδ+ hepatosplenic γδ+ T-cell lymphomas treated on a compassionate basis with dCF. The inverse correlation between CD26 expression and sensitivity to dCF was also demonstrated in a lymphoblastic lymphoma case in which CD26 was expressed on circulating blasts at relapse but not at diagnosis, as well as in two H9 T-cell clones expressing or not expressing CD26 mRNA and protein.

Conclusions: This study corroborates the notion of CD26 as a marker of poor prognosis for T-cell malignancies and delineates a role for CD26 as a predictor of poor response to dCF.
Expression of Transforming Growth Factor β Isoforms and Their Receptors in Malignant Fibrous Histiocytoma of Soft Tissues
Authors: Yamamoto Tetsuji , Akisue Toshihiro , Marui Takashi , Fujita Ikuo , Matsumoto Keiji , Hitora Toshiaki , Kawamoto Teruya , Nagira Keiko , Nakatani Tetsuya , Kurosaka Masahiro
Published: September 2005

Purpose: Transforming growth factor β (TGF-β) is a multifunctional growth factor that variably affects proliferation, differentiation, and extracellular matrix formation. Little information is currently available on the TGF-β expression in malignant fibrous histiocytoma (MFH). The aims of the present study were to investigate the expression of TGF-β isoforms and their receptors in human MFH specimens.

Experimental Design: The expression of TGF isoforms, and TGF-β receptors (TGF-βR1 and -βR2) were immunohistochemically evaluated in 43 paraffin-embedded MFH specimens. Furthermore, the correlation of the TGF-β and receptor expression with tumor proliferative activity assessed by MIB-1 indices was analyzed.

Results: Positive immunoreactivity for TGF-β1, -β2, and -β3 was identified in tumor cells of 42, 40, and 38 of the 43 MFHs, respectively. In each TGF-β isoform immunostaining, the specimens were divided into two groups based on the number of positive tumor cells: those with low (<25%) and those with high (≧25%) immunoreactivity. There were no statistically significant differences in the MIB-1 indices between the two groups. Positive immunoreactivity for TGF-βR1 and -βR2 was identified in tumor cells of 36 and 24 of the MFHs, respectively. The specimens were divided into two groups based on their receptor expression patterns: those with both TGF-βR1- and -βR2-positive immunoreactivity (n = 23), and those with both or either TGF-βR1- and -βR2-negative immunoreactivity (n = 20). The MIB-1 indices in the both-TGF-βR1- and -βR2-positive group were significantly higher than those in the other group (P = 0.0102). There was no significant difference in pulmonary metastasis ratios between the two groups.

Conclusions: These findings strongly suggest an association of the TGF-β ligand/receptor system with a significantly higher MIB-1 index in human MFHs. Investigation of the TGF-βR1 and -βR2 coexpression might be useful in predicting tumor behavior of MFHs.
Pharmacogenomic Analysis of Cytogenetic Response in Chronic Myeloid Leukemia Patients Treated with Imatinib
Authors: McLean Lee Anne , Gathmann Insa , Capdeville Renaud , Polymeropoulos Mihael H. , Dressman Marlene
Published: September 2005

Purpose: To better understand the molecular basis of cytogenetic response in chronic myeloid leukemia patients treated with imatinib, we studied gene expression profiles from a total of 100 patients from a large, multinational Phase III clinical trial (International Randomized Study of IFN-α versus STI571).

Experimental Design: Gene expression data for >12,000 genes were generated from whole blood samples collected at baseline (before imatinib treatment) using Affymetrix oligonucleotide microarrays. Cytogenetic response was determined based on the percentage of Ph+ cells from bone marrow following a median of 13 months of treatment.

Results: A genomic profile of response was developed using a subset of individuals that exhibited the greatest divergence in cytogenetic response; those with complete response (0% Ph+ cells; n = 53) and those with minimal or no response (>65% Ph+ cells; n = 13). A total of 55 genes was identified that were differentially expressed between these two groups. Using a “leave-one-out” strategy, we identified the optimum 31 genes from this list to use as our genomic profile of response. Using this genomic profile, we were able to distinguish between individuals that achieved major cytogenetic response (0–35% Ph+ cells) and those that did not, with a sensitivity of 93.4% (71 of 76 patients), specificity of 58.3% (14 of 24 patients), positive predictive value of 87.7%, and negative predictive value of 73.7%.

Conclusions: Interestingly, many of the genes identified appear to be strongly related to reported mechanisms of BCR-ABL transformation and warrant additional research as potential drug targets. The validity and clinical implications of these results should be explored in future studies.
Chemosensitization of Androgen-Independent Prostate Cancer with Neutral Endopeptidase
Authors: Sumitomo Makoto , Asano Tomohiko , Asakuma Junichi , Asano Takako , Nanus David M. , Hayakawa Masamichi
Published: September 2005

Purpose: We investigated whether neutral endopeptidase (NEP) could augment chemosensitivity to anticancer drugs by promoting protein kinase C (PKC)δ-mediated mitochondrial apoptosis in prostate cancer (PC) cells.

Experimental Design: Human PC cell lines LNCaP and PC-3, and a normal prostate epithelial cell line (PrEC) were used. The protein expression was detected by Western blot analysis, and the protein turnover was determined by pulse-chase assay. Apoptotic ratio was measured by annexin V staining.

Results: Western blot analyses and pulse-chase assays showed that the specific NEP inhibitor CGS24592 decreased PKCδ protein expression by promoting PKCδ protein degradation in NEP-expressing LNCaP cells. Conversely, recombinant NEP (rNEP) increased PKCδ protein expression by delaying PKCδ protein degradation in NEP-negative PC-3 cells. Apoptosis assays showed that rNEP promoted anticancer drug-induced apoptosis in PC-3 cells specifically through PKCδ activity that mediated anticancer drug-induced mitochondrial change such as cytochrome-c release and caspase-9 activation. Of note, rNEP was able to increase PKCδ protein expression predominantly in PC-3 cells rather than in PrEC cells. Treatment with rNEP before subtoxic concentrations of etoposide (0.1 μm) significantly promoted mitochondrial apoptosis compared with only etoposide in PC-3 cells (P < 0.01) but not in PrEC cells.

Conclusions: These results suggest that NEP enzyme activity contributes to anticancer drug-induced PC cell apoptosis dependent on PKCδ-mediated mitochondrial events. More importantly, the combination of NEP with anticancer drugs may be a promising therapeutic modality because rNEP is able to augment chemosensitivity in androgen-independent PC with minimal toxicity in normal tissues.