bluuebunny/crossref_metadata_2025 huggingface dataset:
Biomarkers of Exposure, Effect, and Susceptibility in Workers Exposed to Nitrotoluenes
Authors: Sabbioni Gabriele , Jones Christopher R. , Sepai Ovnair , Hirvonen Ari , Norppa Hannu , Järventaus Hilkka , Glatt Hansruedi , Pomplun Doreen , Yan Huifang , Brooks Lance R. , Warren Sarah H. , DeMarini David M. , Liu Yu-Ying
Published: March 2006

Nitrotoluenes, such as 2-nitrotoluene, 2,4-dinitrotoluene (24DNT), and 26DNT, are carcinogenic in animal experiments. Humans are exposed to such chemicals in the workplace and in the environment. It is therefore important to develop methods to biomonitor people exposed to nitrotoluenes to prevent the potential harmful effects. For the present study, workers exposed to high levels of these chemicals were investigated. The external dose (air levels), the internal dose (urine metabolites), the biologically effective dose [hemoglobin (Hb) adducts and urine mutagenicity], and biological effects (chromosomal aberrations and health effects) were determined. Individual susceptibility was assessed by determining genetic polymorphisms of enzymes assumed to function in nitrotoluene metabolism, namely glutathione S-transferases (GSTM1, GSTT1, GSTP1), N-acetyltransferases (NAT1, NAT2), and sulfotransferases (SULT1A1, SULT1A2). The levels of urinary metabolites did not correlate with the air levels. The urinary mutagenicity levels determined in a subset of workers correlated with the levels of a benzylalcohol metabolite of DNT. The Hb-adducts correlated with the urine metabolites but not with the air levels. The frequency of chromosomal aberrations (gaps included) was increased (P < 0.05) in the exposed workers in comparison with a group of factory controls and correlated with the level of 24DNT Hb-adducts in young subjects (<31 years). The GSTM1-null genotype was significantly more prevalent in the controls than in the exposed group, which probably reflected an elevated susceptibility of the GSTM1-null genotype to adverse health effects of DNT exposure, such as nausea (odds ratio, 8.8; 95% confidence interval, 2.4-32.2). A statistically significant effect was seen for SULT1A2 genotype on a 24DNT Hb-adduct; GSTP1 genotype on a 2,4,6-trinitrotoluene Hb-adduct; and SULT1A1, SULT1A2, NAT1, GSTT1, and GSTP1 genotypes on chromosomal aberrations in the exposed workers. (Cancer Epidemiol Biomarkers Prev 2006;15(3):559–66)
Fruit and Vegetable Consumption in Relation to Pancreatic Cancer Risk: A Prospective Study
Authors: Larsson Susanna C. , Håkansson Niclas , Näslund Ingmar , Bergkvist Leif , Wolk Alicja
Published: March 2006

High consumption of fruits and vegetables has been associated with a lower risk of pancreatic cancer in many case-control studies. However, cohort studies on this relationship are limited and do not support an association. We examined the associations of overall consumption of fruits and vegetables and consumption of certain subgroups of fruits and vegetables with the incidence of pancreatic cancer among 81,922 women and men in the Swedish Mammography Cohort and the Cohort of Swedish Men. Hazard ratios (HR) with 95% confidence intervals (95% CI) were estimated using Cox proportional hazards models. During an average follow-up of 6.8 years (1998-2004), 135 incident pancreatic cancer cases were diagnosed. After adjustment for age and other risk factors for pancreatic cancer, the HRs for the highest compared with the lowest category of intake were 1.13 (95% CI, 0.66-1.94) for total fruits and vegetables, 1.10 (95% CI, 0.64-1.88) for total fruits, and 1.08 (95% CI, 0.63-1.85) for total vegetables. Among specific subgroups of fruits and vegetables, a nonsignificant inverse association was observed with cruciferous vegetable consumption (≥3 servings/wk versus <1 serving/wk: HR, 0.70; 95% CI, 0.43-1.13). Cabbage consumption was associated with a statistically significant lower risk of pancreatic cancer (≥1 serving/wk versus never consumption: HR, 0.62; 95% CI, 0.39-0.99). Findings from this prospective study do not support a relationship of overall fruit and vegetable consumption with pancreatic cancer risk. The association between consumption of cruciferous vegetables and pancreatic cancer risk warrants further investigation. (Cancer Epidemiol Biomarkers Prev 2006;15(2):301–5)
Modeling Exposures for DNA Methylation Profiles
Authors: Siegmund Kimberly D. , Levine A. Joan , Chang Jing , Laird Peter W.
Published: March 2006

We extend the finite mixture model to estimate the association between exposure and latent disease subtype measured by DNA methylation profiles. Estimates from this model are compared with those obtained from the simpler two-phase approach of first clustering the DNA methylation data followed by associating exposure with disease subtype using logistic regression. The two models are fit to data from a study of colorectal adenomas and are compared in a simulation study. Depending on the analytic approach, we obtain different estimates of the odds ratio (OR) and its 95% confidence interval (95% CI) for the association of RBC folate and DNA methylation subtype in colorectal adenomas (OR, 0.31; 95% CI, 0.08-1.26 from the extended finite mixture model; OR, 0.44; 95% CI, 0.15-1.28 from the two-phase approach; n = 58 case subjects). Although our results could be a chance occurrence due to fluctuations from small sample size, we did a simulation study using larger samples and found that differences between the two approaches emerge when there is noise in the cluster analysis. In the naive two-phase approach, the estimate of the OR is biased towards the null, and its SE is underestimated when there is error in the cluster assignment. Estimates from the extended mixture model are unbiased and have the correct SE estimate but may require larger sample sizes for convergence. Thus, when the clusters are not identified with certainty, the extended mixture model is preferred for valid estimation of the OR and CI. (Cancer Epidemiol Biomarkers Prev 2006;15(3):567–72)
Genetic Variation in the Nucleotide Excision Repair Pathway and Bladder Cancer Risk
Authors: García-Closas Montserrat , Malats Núria , Real Francisco X. , Welch Robert , Kogevinas Manolis , Chatterjee Nilanjan , Pfeiffer Ruth , Silverman Debra , Dosemeci Mustafa , Tardón Adonina , Serra Consol , Carrato Alfredo , García-Closas Reina , Castaño-Vinyals Gemma , Chanock Stephen , Yeager Meredith , Rothman Nathaniel
Published: March 2006

Nucleotide excision repair (NER) is critical for protecting against damage from carcinogens in tobacco smoke. We evaluated the influence of common genetic variation in the NER pathway on bladder cancer risk by analyzing 22 single nucleotide polymorphisms (SNP) in seven NER genes (XPC, RAD23B, ERCC1, ERCC2, ERCC4, ERCC5, and ERCC6). Our study population included 1,150 patients with transitional cell carcinoma of the urinary bladder and 1,149 control subjects from Spain. Odds ratios (OR) and 95% confidence intervals (95% CI) were adjusted for age, gender, region, and smoking status. Subjects with the variant genotypes for SNPs in four of the seven genes evaluated had small increases in bladder cancer risk compared to subjects with the homozygous wild-type genotypes: RAD23B IVS5-15A>G (OR, 1.3; 95% CI, 1.1-1.5; P = 0.01), ERCC2 R156R (OR, 1.3; 95% CI, 1.1-1.6; P = 0.006), ERCC1 IVS5+33A>C (OR, 1.2; 95% CI, 1.0-1.5; P = 0.06; Ptrend = 0.04), and ERCC5 M254V (OR, 1.4; 95% CI, 1.0-2.0; P = 0.04). A global test for pathway effects indicated that genetic variation in NER characterized by the 22 SNPs analyzed in this study significantly predicts bladder cancer risk (P = 0.04). Pairwise comparisons suggested that carrying variants in two genes could result in substantial increases in risk. Classification tree analyses suggested the presence of subgroups of individuals defined by smoking and NER genotypes that could have substantial increases in risk. In conclusion, these findings provide support for the influence of genetic variation in NER on bladder cancer risk. A detailed characterization of genetic variation in key NER genes is warranted and might ultimately help identify multiple susceptibility variants that could be responsible for substantial joint increases in risk. (Cancer Epidemiol Biomarkers Prev 2006;15(3):536–42)
Selenium, Apoptosis, and Colorectal Adenomas
Authors: Connelly-Frost Alexandra , Poole Charles , Satia Jessie A. , Kupper Lawrence L. , Millikan Robert C. , Sandler Robert S.
Published: March 2006

Background: Selenium is an essential trace element found in cereals, wheat, dairy products, meat, and fish. This micronutrient may prevent carcinogenesis through several biochemical pathways; one suggested pathway is enhanced apoptosis.

Objectives: The relation between selenium and colorectal adenomas was evaluated because the colorectal adenoma is the established precursor lesion of most colorectal cancers. Apoptosis was a pathway of interest because decreased apoptosis has been associated with an increased prevalence of adenomas. Our objectives were as follows: to investigate the association between (a) selenium and colorectal adenomas and (b) selenium and apoptosis.

Methods: The study population was assembled for the Diet and Health Study III (n = 803), a cross-sectional study conducted at the University of North Carolina Hospital (Chapel Hill, NC). There were 451 participants in the analysis of selenium and adenoma prevalence and 351 participants in the analysis of selenium and apoptosis. Selenium was measured from serum collected at the time of colonoscopy. Apoptosis was measured in biopsies from normal rectal epithelium obtained during the colonoscopy procedure.

Results: Participants in the highest fifth of serum selenium were less likely to have adenomas in comparison with those in the lowest fifth (prevalence ratio, 0.6; 95% confidence interval, 0.4-1.1). Selenium and apoptosis (>2.76 cells per crypt) were not strongly related, but results collectively suggested a roughly inverse association.

Conclusions: High selenium was associated with a reduced prevalence of colorectal adenomas. Apoptosis, however, did not seem to be the mechanism by which selenium was related to adenoma prevalence in our data. (Cancer Epidemiol Biomarkers Prev 2006;15(3):486–93)
Role of Polymorphic Human Cytochrome P450 Enzymes in Estrone Oxidation
Authors: Cribb Alastair E. , Knight M. Joy , Dryer Dagny , Guernsey Judy , Hender Kimberly , Tesch Marvin , Saleh Tarek M.
Published: March 2006

Estrogen and its metabolites are believed to play important roles in breast cancer. The influence of genetic polymorphisms in the enzymes responsible for formation and disposition of estrogen on breast cancer risk may shed light on the importance of estrogen metabolites in this disease. However, for such studies to be valid, it is important to correctly identify the enzymes involved in estrogen bioactivation. Therefore, we assessed the human cytochrome P450–dependent oxidation of estrone using substrate concentrations that more closely approximate the maximum expected concentrations in breast tissue. The in vitro metabolism of estrone by recombinant human cytochrome P450 enzymes and human liver microsomes was studied. The formation of estrone metabolites (2-hydroxyestrone, 4-hydroxyestrone, and 16α-hydroxyestrone) was monitored by high-performance liquid chromatography. 2-Hydroxyestrone formation was catalyzed predominantly by CYP1A2, CYP1A1, and CYP1B1 enzymes; 4-hydroxyestrone formation was catalyzed predominantly by CYP1B1, CYP1A2, and CYP1A1 enzymes; and 16α-hydroxyestrone formation was catalyzed predominantly by CYP2C19, CYP1A1, and CYP3A5. This study confirms the important role of members of the CYP1 family in the 2-hydroxylation and 4-hydroxylation of estrone, but the enzymes identified as responsible for the 16α-hydroxylation of estrone are different from those previously identified. The relative importance of these enzymes in vivo would depend on the specific tissue expression of the enzymes. These enzymes are all known to be genetically variant in the human population, and additional studies to assess the role CYP1A2, CYP2C19, and CYP3A5 in breast cancer risk are indicated. (Cancer Epidemiol Biomarkers Prev 2006;15(3):551–8)
Prostacyclin Synthase and Arachidonate 5-Lipoxygenase Polymorphisms and Risk of Colorectal Polyps
Authors: Poole Elizabeth M. , Bigler Jeannette , Whitton John , Sibert Justin G. , Potter John D. , Ulrich Cornelia M.
Published: March 2006

Prostacyclin synthase (PGIS) and arachidonate 5-lipoxygenase (ALOX5) are enzymes relevant to prostaglandin and leukotriene synthesis, both important pathways for colon cancer risk. We hypothesized that genetic variation altering the function of these enzymes would modify risk of colorectal polyps. In a Minnesota-based case-control study of adenomatous (n = 517) or hyperplastic (n = 192) polyps versus polyp-free controls (n = 618), we investigated the role of promoter repeat polymorphisms in PGIS and ALOX5 as well as ALOX5 −1700 G>A. Having fewer than six repeats on both PGIS alleles (<6R/<6R) was associated with an increased risk of adenomas compared with the 6R/6R (wild-type) genotype (OR, 1.90; 95% CI, 1.09-3.30). Having more repeats (>6R/≥6R) reduced risk (OR, 0.73; 95% CI, 0.40-1.35; Ptrend = 0.03). In allele-based analyses, fewer repeats were associated with a modestly increased risk of adenomas and perhaps hyperplastic polyps. There were no risk differences for either the ALOX5 VNTR or −1700 G>A polymorphisms. Associations with regular use of aspirin or other nonsteroidal anti-inflammatory drugs (NSAIDs) differed by PGIS genotype. Among individuals with at least one wild-type allele, NSAID use was associated with a decreased risk; however, those with fewer PGIS repeats (<6R/<6R) did not benefit (Pinteraction = 0.06). There was also evidence of an interaction between the COX-2 −765 G>C and ALOX5 −1700 G>A genotypes (Pinteraction = 0.07). The PGIS promoter polymorphism may affect risk of colorectal polyps and modify the effects of NSAID use on polyp risk. A more comprehensive investigation of genetic variability in prostaglandin synthesis in relation to risk of colorectal neoplasia and NSAID pharmacogenetics is warranted. (Cancer Epidemiol Biomarkers Prev 2006;15(3):502–8)
Measurements of Breast Density: No Ratio for a Ratio
Authors: Haars Gerco , van Noord Paulus A.H. , van Gils Carla H. , Grobbee Diederick E. , Peeters Petra H.M.
Published: March 2006

Breast density, as visible on mammograms, is generally assessed as the occupied percentage of the breast and is a risk factor for breast cancer. Various studies have looked into the causation and alteration of relative density but the relation of a determinant with a relative measure does not allow a direct etiologic interpretation. It was our goal to compare the effects of known determinants on relative density and the absolute amounts of dense and nondense tissues. We measured the absolute and relative densities in a population of 418 postmenopausal women participating in a breast cancer screening program. The occupied surface area was calculated after manually tracing the contours of the tissues on digitized mammograms. Information on determinants was available through physical examination and questionnaires. Data were analyzed by multivariate linear regression. Age and parity were found to decrease the amount of dense tissue and the ages at menarche and menopause were found to increase it (R2 = 13%). The amount of nondense tissue was increased by higher body mass index (BMI), age, and parity (R2 = 43%). Relative density was affected by a combination of these factors (R2 = 29%) with directionalities of effects that are comparable to those of dense tissue. However, the magnitudes of these effects were the resultant of the effects on dense and nondense tissues. The influence of BMI on relative density was completely due to an effect on nondense tissue. Although relative density is a relevant prognostic factor, inferences about the etiology of breast density should be made on the basis of absolute measures.
Activation of Stat3 in Primary Tumors from High-Risk Breast Cancer Patients Is Associated with Elevated Levels of Activated Src and Survivin Expression
Authors: Diaz Nills , Minton Susan , Cox Charles , Bowman Tammy , Gritsko Tanya , Garcia Roy , Eweis Ibrahim , Wloch Marek , Livingston Sandy , Seijo Ed , Cantor Alan , Lee Ji-Hyun , Beam Craig A. , Sullivan Daniel , Jove Richard , Muro-Cacho Carlos A.
Published: March 2006

Purpose: Constitutive activation of signal transducer and activator of transcription 3 (Stat3) protein has been observed in a wide variety of tumors, including breast cancer, and contributes to oncogenesis at least in part by prevention of apoptosis. In a study of 45 patients with high-risk breast cancer enrolled in a phase II neoadjuvant chemotherapy trial with docetaxel and doxorubicin, we evaluated the levels of Stat3 activation and potentially associated molecular biomarkers in invasive breast carcinoma compared with matched nonneoplastic tissues.

Experimental Design: Using immunohistochemistry and image analysis, we quantified the levels of phospho-Stat3 (pY-Stat3), phospho-Src (pY-Src), epidermal growth factor receptor, HER2/neu, Ki-67, estrogen receptor, Bcl-2, Bcl-xL, Survivin, and apoptosis in formalin-fixed, paraffin-embedded sections from invasive carcinomas and their paired nonneoplastic parenchyma. The levels of molecular biomarkers in nonneoplastic and tumor tissues were analyzed as continuous variables for statistically significant correlations.

Results: Levels of activated pY-Stat3 and pY-Src measured by immunohistochemistry were significantly higher in invasive carcinoma than in nonneoplastic tissue (P < 0.001). In tumors, elevated levels of pY-Stat3 correlated with those of pY-Src and Survivin. Levels of pY-Stat3 were higher in partial pathologic responders than in complete pathologic responders. In partial pathologic responders, pY-Stat3 levels correlated with Survivin expression.

Conclusions: Our findings suggest important roles for elevated activities of Stat3 and Src, as well as Survivin expression, in malignant progression of breast cancer. Furthermore, elevated Stat3 activity correlates inversely with complete pathologic response. These findings suggest that specific Stat3 or Src inhibitors could offer clinical benefits to patients with breast cancer.
Persistent Activation of Stat3 Signaling Induces Survivin Gene Expression and Confers Resistance to Apoptosis in Human Breast Cancer Cells
Authors: Gritsko Tanya , Williams Ann , Turkson James , Kaneko Satoshi , Bowman Tammy , Huang Mei , Nam Sangkil , Eweis Ibrahim , Diaz Nils , Sullivan Daniel , Yoder Sean , Enkemann Steve , Eschrich Steven , Lee Ji-Hyun , Beam Craig A. , Cheng Jin , Minton Susan , Muro-Cacho Carlos A. , Jove Richard
Published: March 2006

Purpose: Signal transducer and activator of transcription 3 (Stat3) protein is persistently activated in breast cancer and promotes tumor cell survival. To gain a better understanding of the role of constitutive Stat3 signaling in breast cancer progression, we evaluated the expression profile of potential Stat3-regulated genes that may confer resistance to apoptosis.

Experimental Design: Stat3 signaling was blocked with antisense oligonucleotides in human MDA-MB-435s breast cancer cells and Affymetrix GeneChip microarray analysis was done. The candidate Stat3 target gene Survivin was further evaluated in molecular assays using cultured breast cancer cells and immunohistochemistry of breast tumor specimens.

Results: Survivin, a member of the inhibitor of apoptosis protein family, was identified as a potential Stat3-regulated gene by microarray analysis. This was confirmed in Survivin gene promoter studies and chromatin immunoprecipitation assays showing that Stat3 directly binds to and regulates the Survivin promoter. Furthermore, direct inhibition of Stat3 signaling blocked the expression of Survivin protein and induced apoptosis in breast cancer cells. Direct inhibition of Survivin expression also induced apoptosis. Increased Survivin protein expression correlates significantly (P = 0.001) with elevated Stat3 activity in primary breast tumor specimens from high-risk patients who were resistant to chemotherapy treatment.

Conclusions: We identify Survivin as a direct downstream target gene of Stat3 in human breast cancer cells that is critical for their survival in culture. Our findings suggest that activated Stat3 signaling contributes to breast cancer progression and resistance to chemotherapy by, at least in part, inducing expression of the antiapoptotic protein, Survivin.
Gene Expression Profiles of B-lineage Adult Acute Lymphocytic Leukemia Reveal Genetic Patterns that Identify Lineage Derivation and Distinct Mechanisms of Transformation
Authors: Chiaretti Sabina , Li Xiaochun , Gentleman Robert , Vitale Antonella , Wang Kathy S. , Mandelli Franco , Foà Robin , Ritz Jerome
Published: March 2006

Purpose: To characterize gene expression signatures in acute lymphocytic leukemia (ALL) cells associated with known genotypic abnormalities in adult patients.

Experimental Design: Gene expression profiles from 128 adult patients with newly diagnosed ALL were characterized using high-density oligonucleotide microarrays. All patients were enrolled in the Italian GIMEMA multicenter clinical trial 0496 and samples had >90% leukemic cells. Uniform phenotypic, cytogenetic, and molecular data were also available for all cases.

Results: T-lineage ALL was characterized by a homogeneous gene expression pattern, whereas several subgroups of B-lineage ALL were evident. Within B-lineage ALL, distinct signatures were associated with ALL1/AF4 and E2A/PBX1 gene rearrangements. Expression profiles associated with ALL1/AF4 and E2A/PBX1 are similar in adults and children. BCR/ABL+ gene expression pattern was more heterogeneous and was most similar to ALL without known molecular rearrangements. We also identified a set of 83 genes that were highly expressed in leukemia blasts from patients without known molecular abnormalities who subsequently relapsed following therapy. Supervised analysis of kinase genes revealed a high-level FLT3 expression in a subset of cases without molecular rearrangements. Two other kinases (PRKCB1 and DDR1) were highly expressed in cases without molecular rearrangements, as well as in BCR/ABL-positive ALL.

Conclusions: Genomic signatures are associated with phenotypically and molecularly well defined subgroups of adult ALL. Genomic profiling also identifies genes associated with poor outcome in cases without molecular aberrations and specific genes that may be new therapeutic targets in adult ALL.
Effect of the St. John's Wort Constituent Hyperforin on Docetaxel Metabolism by Human Hepatocyte Cultures
Authors: Komoroski Bernard J. , Parise Robert A. , Egorin Merrill J. , Strom Stephen C. , Venkataramanan Raman
Published: March 2006

Background and Purpose: St. John's wort is a commonly used herbal medication that increases cytochrome P450 3A (CYP3A) activity. Because docetaxel is inactivated by CYP3A, we studied the effects of the St. John's wort constituent hyperforin on docetaxel metabolism in a human hepatocyte model.

Experimental Design: Hepatocytes, isolated from three donor livers, were exposed to hyperforin (0.1, 0.5, or 1.5 μmol/L) or rifampin (10 μmol/L) for 48 hours. After 48 hours, hyperforin- or rifampin-containing medium was replaced with medium containing 100 μmol/L docetaxel. After 1 hour, docetaxel metabolism was characterized by liquid chromatography-tandem mass spectrometry. Subsequent incubations characterized the specific cytochrome P450s that produced the docetaxel metabolites observed in hepatocyte incubations.

Results: Rifampin induced docetaxel metabolism 6.8- to 32-fold above docetaxel metabolism in control cultures. Hyperforin induced docetaxel metabolism in all three hepatocyte preparations. Hyperforin induction was dose-dependent and, at maximum, was 2.6- to 7-fold greater than that in controls. Docetaxel metabolites identified in rifampin- and hyperforin-treated hepatocyte preparations included the previously described tert-butyl–hydroxylated metabolite and two previously unidentified metabolites involving hydroxylation on the baccatin ring. CYP3A4 produced the tert-butyl–hydroxylated metabolite and the two ring-hydroxylated metabolites. CYP2C8 produced one of the newly described ring-hydroxylated metabolites.

Conclusions: Exposure to the St. John's wort constituent hyperforin induces docetaxel metabolism in vitro. This implies that subtherapeutic docetaxel concentrations may result when docetaxel is administered to patients using St. John's wort on a chronic basis. The results also show induction of previously undescribed metabolic pathways for docetaxel, one of which may be analogous to the known 6-α-hydroxylation of paclitaxel by CYP2C8.
Absence of Adverse Effects in Cynomolgus Macaques Treated with CNTO 95, a Fully Human Anti-αv Integrin Monoclonal Antibody, Despite Widespread Tissue Binding
Authors: Martin Pauline L. , Jiao Qun , Cornacoff Joel , Hall William , Saville Bradley , Nemeth Jeffrey A. , Schantz Allen , Mata Marielena , Jang Haishan , Fasanmade Adedigbo A. , Anderson Lisa , Graham Martin A. , Davis Hugh M. , Treacy George
Published: March 2006

Purpose: CNTO 95 is a fully human anti-αv integrin monoclonal antibody that inhibits macaque and rodent angiogenesis and inhibits human tumor growth in rodents. The purpose of these studies was to evaluate the preclinical safety of long-term administration of CNTO 95 in cynomolgus macaques.

Experimental Design: The in vitro binding profiles of CNTO 95 to human and macaque tissues and the in vivo binding to macaque tissues was evaluated by immunohistochemistry. The preclinical safety of CNTO 95 (10 and 50 mg/kg, i.v.) was evaluated in macaques treated once per week for up to 6 months. Safety was evaluated by clinical observations, ophthalmic and physical examinations (including heart rate, blood pressure, and electrocardiogram), clinical pathology (including coagulation parameters), and comprehensive anatomic pathology. The effect of CNTO 95 (50 mg/kg, i.v.) on incisional wound healing was evaluated in macaques.

Results: The tissue binding studies showed that CNTO 95 bound with mild to moderate intensity to macaque and human endothelial cells, epithelial cells, and vascular smooth muscle cells in most normal tissues examined. CNTO 95 showed strong to intense staining to the positive control tissue, human placenta. Despite the widespread binding to normal tissues, treatment of cynomolgus macaques with CNTO 95 produced no signs of toxicity and no histopathologic changes in any of the tissues examined (including ovaries and bone growth plates). CNTO 95 did not impair wound healing.

Conclusion: These studies show that CNTO 95 is safe and, unlike some other angiogenesis inhibitors, does not seem to inhibit normal physiologic angiogenesis.
Evaluation of a Panel of Molecular Markers for the Diagnosis of Malignant Serous Effusions
Authors: Passebosc-Faure Karine , Li Guorong , Lambert Claude , Cottier Michèle , Gentil-Perret Anne , Fournel Pierre , Pérol Maurice , Genin Christian
Published: March 2006

Purpose: Our main goal was to evaluate a panel of molecular markers for the detection of cancer cells in serous effusions and to determine their value as an adjunctive reverse transcription-PCR (RT-PCR) test to cytologic examination.

Experimental Design: One hundred fourteen serous effusions from 71 patients with tumors and 43 patients with benign diseases were subjected to RT-PCR for expression of carcinoembryonic antigen (CEA), epithelial cell adhesion molecule (Ep-CAM), E-cadherin, mammaglobin, mucin 1 (MUC1) isoforms MUC1/REP, MUC1/Y, and MUC1/Z, calretinin, and Wilms' tumor 1 susceptibility gene.

Results: CEA, Ep-CAM, E-cadherin, and mammaglobin were specifically expressed in malignant effusions. The sensitivity of RT-PCR in cytologically negative malignant effusions was 63.1% combining CEA and Ep-CAM (with 100% specificity) and reached 78.9% adding MUC1/Y or MUC1/Z (with 93% specificity). In the whole population of effusions, the combination of cytology with RT-PCR of CEA and Ep-CAM yielded a 90.1% sensitivity, a specificity and a positive predictive value of 100%, and a 86% negative predictive value for malignancy. Adding MUC1/Y or MUC1/Z to the panel, the sensitivity was 94.5% with 93% specificity, 95.7% PPV, and 90.9% negative predictive value. Moreover, CEA and mammaglobin were specifically expressed in epithelial malignancies, and mammaglobin was mainly expressed in effusions from breast carcinoma (97.3% of specificity).

Conclusions: A combination of cytology and RT-PCR analysis of CEA and Ep-CAM significantly improved the detection sensitivity of tumor cells in serous effusions. RT-PCR analysis of CEA, Ep-CAM, and mammaglobin in serous effusions could be a beneficial adjunct to cytology for the diagnosis of malignancy.
Thomsen-Friedenreich and Tn Antigens in Nipple Fluid: Carbohydrate Biomarkers for Breast Cancer Detection
Authors: Kumar Senthil R. , Sauter Edward R. , Quinn Thomas P. , Deutscher Susan L.
Published: March 2006

Purpose: Novel biomarkers would facilitate early and accurate diagnosis of breast cancer. The Thomsen-Freidenreich (TF) and Tn antigens are aberrantly glycosylated carbohydrate cancer-associated antigens found in ∼80% of adenocarcinomas. Both TF and Tn are expressed on cell-surface glycoproteins and glycolipids. Nipple aspirate fluid (NAF) is concentrated in secreted proteins and lipids from cells that give rise to cancer. The objective of this study was to determine if NAF from breasts with cancer contains elevated levels of TF and Tn compared with NAF from normal breasts. A sensitive and specific antigen capture immunoassay for TF and Tn detection in NAF was developed for this purpose.

Experimental Design: Fifty NAF samples, 25 from breasts with cancer and 25 from normal breasts, were examined. Antigen capture immunoassays were done on the samples using monoclonal antibodies that specifically recognized either TF or Tn antigen in NAF. These antibodies captured serially diluted NAF samples, and the concentration of TF or Tn was determined by comparing absorbance values against a standard curve generated from standard sources of TF or Tn.

Results: TF and Tn were detected in 19 of 25 and 20 of 25 NAF samples from breasts with cancer, respectively, compared with 0 of 25 and 1 of 25 NAF samples from breasts without cancer (P < 0.001 for both TF and Tn). In 92% of the cancerous breast NAF samples tested, either TF or Tn was found.

Conclusions: Simultaneous measurement of TF and Tn in NAF may facilitate the noninvasive detection of breast cancer and warrants further study.
Phosphatidylinostol 3-Kinase Mediates Angiogenesis and Vascular Permeability Associated with Ovarian Carcinoma
Authors: Hu Limin , Hofmann Judith , Jaffe Robert B.
Published: March 2006

Purpose: To assess the role of phosphatidylinositol-3 kinase (PI3K) inhibition in vascular permeability, angiogenesis, and vascular remodeling in tumor vessels and peritoneal lining in an athymic mouse model of i.p. human ovarian carcinoma.

Experimental Design: Mice were inoculated i.p. with cells from the human ovarian cancer cell line, OVCAR-3. Fourteen days after inoculation, mice were treated with or without the PI3K inhibitor LY294002, 3 days weekly for 4 weeks. At the end of the experiment, some mice were anesthetized and injected via the tail vein with FITC-labeled lycopersicon lectin and perfused through the aorta before sacrifice. The peritoneal wall and tumor from all mice were removed and embedded in 10% agarose. Tumor sections were visualized by fluorescence microscopy.

Results: Ascites in the LY294002-treated group (0.69 ± 0.27 mL) was reduced by 72.4% compared with the control group (2.5 ± 1.2 mL). Tumor burden in the LY294002-treated group (0.62 ± 0.32 g) was reduced by 47.3% compared with the control group (1.18 ± 0.41 g). LY294002 inhibited peritoneal and tumor vascularization resulting in numerous leaky, irregular, tortuous vessels in scant, straight, relatively impermeable vessels.

Conclusions: The data indicate that LY294002 inhibits ascites formation in our mouse model of human ovarian cancer by inhibiting tumor and peritoneal neovascularization as well as vascular permeability. The data also show that LY294002 directly inhibits vascular endothelial growth factor (VEGF) protein expression and release from ovarian carcinoma and suggest that LY294002 blocks the VEGF signaling pathway involved in angiogenesis and vascular permeability.
A Physiologic Imaging Pilot Study of Breast Cancer Treated with AZD2171
Authors: Miller Kathy D. , Miller Michael , Mehrotra Sanjana , Agarwal Beamon , Mock Bruce H. , Zheng Qi-Huang , Badve Sunil , Hutchins Gary D. , Sledge George W.
Published: March 2006

Background: This pilot study combined physiologic imaging, microcomputed tomography, and histologic tumor evaluation with a xenograft model of breast cancer to identify surrogates likely to correlate with response to AZD2171, an inhibitor of the vascular endothelial growth factor (VEGF) receptor tyrosine kinases.

Experimental Design: MCF-7 cells transfected with vector (MCF-7neo) or VEGF (MCFVEGF) were implanted in the right and left mammary fat pads of 75 athymic mice. Treatment with AZD2171 (5 mg/kg/d) or vehicle control was initiated once tumors were established. Positron emission tomography with [11C]carbon monoxide to measure blood volume, [18F]fluoromethane to measure perfusion, and [18F]fluorodeoxyglucose to measure glucose utilization was done at baseline, and after 24 hours, 72 hours, and 4 weeks of treatment. After imaging, tumors were analyzed for microvessel density, proliferation, and VEGF expression.

Results: AZD2171 induced significant inhibition of tumor growth in established MCF-7neo xenografts and regression of established MCF-7VEGF xenografts. An acute decrease in blood flow was detected in MCF-7VEGF tumors at 24 hours (P = 0.05). Tumor blood volume was increased in the MCF-7VEGF tumors but correlated with tumor size; blood volume did not change with AZD2171 therapy. Glucose utilization correlated with tumor size and did not change with acute or chronic AZD2171 therapy. Unlike blood flow and blood volume, glucose utilization was similar in MCF-7neo and MCF-7VEGF tumors. Microvessel density and proliferation acutely decreased in MCF-7VEGF tumors but returned to baseline during chronic therapy.

Conclusions: [18F]Fluoromethane imaging may be a useful surrogate for biological activity of AZD2171 with changes identified within 24 hours of starting therapy.
Phase I Clinical Trial of a HER-2/neu Peptide (E75) Vaccine for the Prevention of Prostate-Specific Antigen Recurrence in High-Risk Prostate Cancer Patients
Authors: Hueman Matthew T. , Dehqanzada Zia A. , Novak Thomas E. , Gurney Jennifer M. , Woll Michael M. , Ryan Gayle B. , Storrer Catherine E. , Fisher Christine , McLeod David G. , Ioannides Constantine G. , Ponniah Sathibalan , Peoples George E.
Published: March 2006

Purpose: The E75 peptide is an immunogenic peptide from the HER-2/neu protein that is substantially expressed in prostate cancer. We are conducting a clinical trial of an E75/granulocyte macrophage colony-stimulating factor vaccine to prevent post-prostatectomy prostate-specific antigen (PSA) recurrences in high-risk prostate cancer (HRPC) patients.

Experimental Design: Prostate cancer patients at high risk for recurrence were prospectively evaluated and identified by the validated Center for Prostate Disease Research (CPDR)/CaPSURE high-risk equation. From these high-risk equation patients, 27 HER-2/neu-expressing prostate cancer patients were enrolled. HLA-A2+ patients (n = 17) were vaccinated, whereas HLA-A2− patients (n = 10) were followed as clinical controls. Local/systemic toxicities, immunologic responses, and time to recurrence were measured.

Results: This vaccine is safe with only minor toxicities observed. Additionally, the vaccine is immunogenic with all patients showing both in vivo and in vitro phenotypic and functional immune responses, although variable. HLA-A2+ patients were found to have larger tumors, higher postoperative Gleason scores, and more high-risk CPDR scores than HLA-A2− patients. Despite these differences, disease-free survival was not different between the vaccinated HLA-A2+ patients and the HLA-A2− controls at a median follow up of 23 months. Three of the four vaccinated patients that recurred had rising PSAs at the initiation of the trial. Ex vivo phenotypic assays were predictive of recurrences and correlated in general with functional assays.

Conclusions: The E75 vaccine strategy is safe and effective in eliciting an immune response against the HER-2/neu protein in HRPC patients and may be useful as a preventive strategy against disease recurrence. Vaccination in response to a rising PSA may be too late.
A Phase II Trial of Pemetrexed in Advanced Breast Cancer: Clinical Response and Association with Molecular Target Expression
Authors: Gomez Henry L. , Santillana Sergio L. , Vallejos Carlos S. , Velarde Raul , Sanchez Juvenal , Wang Xinpeng , Bauer Nancy L. , Hockett Richard D. , Chen Victor J. , Niyikiza Clet , Hanauske Axel R.
Published: March 2006

Purpose: This phase II trial of pemetrexed explored potential correlations between treatment outcome (antitumor activity) and molecular target expression.

Experimental Design: Chemonaïve patients with advanced breast cancer received up to three cycles of pemetrexed 500 mg/m2 (10-minute i.v. infusion) on day 1 of a 21-day cycle, with folic acid and vitamin B12 supplementation. Tumors were surgically removed after the last cycle of pemetrexed as clinically indicated. Biopsies were taken at baseline, 24 hours after infusion in cycle 1, and after cycle 3.

Results: Sixty-one women (median age, 46 years; range, 32-72 years) were treated and were evaluable for response. Objective response rate was 31%. Simple logistic regression suggested a potential relationship between mRNA expression of thymidylate synthase (TS) and pemetrexed response (P = 0.103). Based on threshold analysis, patients with “low” baseline TS (≤71) were more likely to respond to pemetrexed than patients with “high” baseline TS (>71). Expression of baseline dihydrofolate reductase and glycinamide ribonucleotide formyl transferase tended to be higher in responders but this association was not significant (P > 0.311). TS expression increased significantly between baseline and biopsy 2 (P = 0.004) and dropped to near baseline levels at biopsy 3. Conversely, dihydrofolate reductase and glycinamide ribonucleotide formyl transferase decreased after pemetrexed chemotherapy.

Conclusions: Our results suggest a potential association between “low” pretreatment TS expression levels and response to pemetrexed chemotherapy. Future trials examining expression levels of other genes important to the folate pathway and/or breast cancer may identify a more robust multigene profile that can better predict response to this novel antifolate.
Complete Tumor Response Following Intratumoral 32P BioSilicon on Human Hepatocellular and Pancreatic Carcinoma Xenografts in Nude Mice
Authors: Zhang Kai , Loong Susan L.E. , Connor Steve , Yu Sidney W.K. , Tan Soo-Yong , Ng Robert T.H. , Lee Khai Mun , Canham Leigh , Chow Pierce K.H.
Published: March 2006

Purpose: 32P BioSilicon is a new, implantable, radiological medical device that comprises particles of highly pure silicon encapsulating 32phosphorus (32P) for the treatment of unresectable solid tumors. Prior to administration, the device particles are suspended in a formulant which provides an even suspension of the intended dose for implantation. The primary objective of this animal trial study was to investigate the effects of intratumoral injection of 32P BioSilicon on human hepatocellular (HepG2) and pancreatic carcinoma (2119) xenografts implanted in nude mice (BALB/c). A secondary objective was the histopathologic examination of the tumor foci and surrounding tissue during the study.

Methods: Cultured human carcinoma cells (HepG2 and 2119) were injected s.c. into the gluteal region of nude mice. When the implanted tumors were ∼1 cm in diameter, 32P BioSilicon (0.5, 1.0, and 2.0 MBq) or formulant was injected into the tumors. Implanted tumor size was measured once a week for 10 weeks. At study termination, the tumor and surrounding normal tissue were collected and fixed in 10% formalin and processed for histopathologic analysis.

Results: 32P BioSilicon produced a reduction in HepG2 tumor volume when compared with formulant control, and complete response was observed among tumors in the 1.0 and 2.0 MBq treatment groups after week 8. There was also significant reduction in 2119 tumor volume in all treated groups, with the complete response rate of 67% in the 2.0 MBq group.

Conclusion: 32P BioSilicon suppressed the growth of both human hepatocellular and pancreatic carcinoma xenografts implanted in nude mice and complete responses were also observed in tumors at higher radiation doses.
Reversal of Hypermethylation and Reactivation of p16INK4a, RARβ, and MGMT Genes by Genistein and Other Isoflavones from Soy
Authors: Fang Ming Zhu , Chen Dapeng , Sun Yi , Jin Zhe , Christman Judith K. , Yang Chung S.
Published: March 2006

Purpose: We have previously shown the reactivation of some methylation-silenced genes in cancer cells by (−)-epigallocatechin-3-gallate, the major polyphenol from green tea. To determine whether other polyphenolic compounds have similar activities, we studied the effects of soy isoflavones on DNA methylation.

Experimental Design: Enzyme assay was used to determine the inhibitory effect of genistein on DNA methyltransferase activity in nuclear extracts and purified recombinant enzyme. Methylation-specific PCR and quantitative real-time PCR were employed to examine the DNA methylation and gene expression status of retinoic acid receptor β (RARβ), p16INK4a, and O6-methylguanine methyltransferase (MGMT) in KYSE 510 esophageal squamous cell carcinoma cells treated with genistein alone or in combination with trichostatin, sulforaphane, or 2′-deoxy-5-aza-cytidine (5-aza-dCyd).

Results: Genistein (2-20 μmol/L) reversed DNA hypermethylation and reactivated RARβ, p16INK4a, and MGMT in KYSE 510 cells. Genistein also inhibited cell growth at these concentrations. Reversal of DNA hypermethylation and reactivation of RARβ by genistein were also observed in KYSE 150 cells and prostate cancer LNCaP and PC3 cells. Genistein (20-50 μmol/L) dose-dependently inhibited DNA methyltransferase activity, showing substrate- and methyl donor–dependent inhibition. Biochanin A and daidzein were less effective in inhibiting DNA methyltransferase activity, in reactivating RARβ, and in inhibiting cancer cell growth. In combination with trichostatin, sulforaphane, or 5-aza-dCyd, genistein enhanced reactivation of these genes and inhibition of cell growth.

Conclusions: These results indicate that genistein and related soy isoflavones reactivate methylation-silenced genes, partially through a direct inhibition of DNA methyltransferase, which may contribute to the chemopreventive activity of dietary isoflavones.
Meta-analysis of the p53 Mutation Database for Mutant p53 Biological Activity Reveals a Methodologic Bias in Mutation Detection
Authors: Soussi Thierry , Asselain Bernard , Hamroun Dalil , Kato Shunsuke , Ishioka Chikashi , Claustres Mireille , Béroud Christophe
Published: March 2006

Purpose: Analyses of the pattern of p53 mutations have been essential for epidemiologic studies linking carcinogen exposure and cancer. We were concerned by the inclusion of dubious reports in the p53 databases that could lead to controversial analysis prejudicial to the scientific community.

Experimental Design: We used the universal mutation database p53 database (21,717 mutations) combined with a new p53 mutant activity database (2,300 mutants) to perform functional analysis of 1,992 publications reporting p53 alterations. This analysis was done using a statistical approach similar to that of clinical meta-analyses.

Results: This analysis reveals that some reports of infrequent mutations are associated with almost normal activities of p53 proteins. These particular mutations are frequently found in studies reporting multiple mutations in one tumor, silent mutations, or lacking mutation hotspots. These reports are often associated with particular methodologies, such as nested PCR, for which key controls are not satisfactory.

Conclusions: We show the importance of accurate functional analysis before inferring any genetic variation. The quality of the p53 databases is essential in order to prevent erroneous analysis and/or conclusions. The availability of functional data from our new p53 web site (http://p53.free.fr and http://www.umd.be:2072/) will allow functional prescreening to identify potential artifactual data.
Intermittent Exposure of Primitive Quiescent Chronic Myeloid Leukemia Cells to Granulocyte-Colony Stimulating Factor In vitro Promotes their Elimination by Imatinib Mesylate
Authors: Jørgensen Heather G. , Copland Mhairi , Allan Elaine K. , Jiang Xiaoyan , Eaves Allen , Eaves Connie , Holyoake Tessa L.
Published: March 2006

Purpose: Primitive quiescent chronic myeloid leukemia (CML) cells are biologically resistant to imatinib mesylate, an inhibitor of the p210BCR-ABL kinase. The present study was designed to investigate whether either continuous or intermittent exposure of these cells to granulocyte-colony stimulating factor (G-CSF) in vitro can overcome this limitation to the effectiveness of imatinib mesylate therapy.

Experimental Design: CD34+ leukemic cells were isolated from six newly diagnosed chronic phase CML patients and cultured for 12 days in serum-free medium with or without G-CSF and/or imatinib mesylate present either continuously or intermittently (three cycles of G-CSF for 0, 1, or 4 days ± imatinib mesylate for 0, 3, or 4 days). Every 4 days, the number of residual undivided viable cells and the total number of viable cells present were measured.

Results: Intermittent but not continuous exposure to G-CSF significantly accelerated the disappearance in vitro of initially quiescent CD34+ CML cells. This resulted in 3- and 5-fold fewer of these cells remaining after 8 and 12 days, respectively, relative to continuous imatinib mesylate alone (P < 0.04). Cultures containing imatinib mesylate and intermittently added G-CSF also showed the greatest reduction in the total number of cells present after 12 days (5-fold more than imatinib mesylate alone).

Conclusion: Intermittent exposure to G-CSF can enhance the effect of imatinib mesylate on CML cells by specifically targeting the primitive quiescent leukemic elements. A protocol for treating chronic-phase CML patients with imatinib mesylate that incorporates intermittent G-CSF exposure may offer a novel strategy for obtaining improved responses in vivo.