bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunoquantitative Real-Time PCR for Detection and Quantification of
Staphylococcus aureus
Enterotoxin B in Foods
Authors: Rajkovic Andreja , Moualij Benaissa El , Uyttendaele Mieke , Brolet Philippe , Zorzi Willy , Heinen Ernst , Foubert Ellen , Debevere Johan
Published: October 2006

A real-time immunoquantitative PCR (iqPCR) method for detection of
Staphylococcus aureus
enterotoxin B (SEB) was developed and evaluated using both pure cultures and foods. The assay consisted of immunocapture of SEB and real-time PCR amplification of the DNA probe linked to the detection antibody. iqPCR was compared to an in-house enzyme-linked immunosorbent assay (ELISA) using the same couple of capture-detection antibodies and to commercial kits for detection of
S. aureus
enterotoxins (SE). The iqPCR was approximately 1,000 times more sensitive (<10 pg ml
−1
) than the in-house ELISA and had a dynamic range of approximately 10 pg ml
−1
to approximately 30,000 pg ml
−1
. iqPCR was not inhibited by any of the foods tested and was able to detect SEB present in these foods. No cross-reactivity with SE other than SEB was observed. Application of iqPCR for detection of SEB in cultures of
S. aureus
revealed the onset of SEB production after 4 h of incubation at 22, 37, and 42°C, which was in the first half of the exponential growth phase. The total amounts of SEB produced by the two strains tested were larger at 42°C than at 37°C and were strain dependent.
Use of a Multiantigen Detection Algorithm for Diagnosis of Kaposi's Sarcoma-Associated Herpesvirus Infection
Authors: Laney A. Scott , Peters Jonathan S. , Manzi Susan M. , Kingsley Lawrence A. , Chang Yuan , Moore Patrick S.
Published: October 2006

The ability to readily and accurately diagnose Kaposi's sarcoma-associated herpesvirus (KSHV, or human herpesvirus 8) infection in individuals remains a demanding task. Among the available diagnostic methods, sensitivities and specificities range widely, and many are inadequate for large-scale screening studies. We examined a serological algorithm for detecting KSHV in human sera having high sensitivity and specificity. This method uses previously described open reading frame (ORF) K8.1 and ORF65 peptide-based enzyme-linked immunosorbent assays and a novel purified recombinant full-length LANA1 protein. We generated two multiantigen algorithms: one that maximized sensitivity and one that maximized specificity. These serological algorithms were then used to evaluate seroprevalence rates among populations of clinical and epidemiological importance. The serological algorithms yielded sensitivities of 96% and 93% and specificities of 94% and 98% for the more sensitive and specific algorithms, respectively. Among kidney donors, seroprevalence was low, 4.0% (2/50), and similar to that of blood donors (
P
= 0.46; odds ratio [OR], 1.4; confidence interval [CI], 0.14 to 7.9) using the highly specific algorithm. Using the sensitive algorithm, 8.0% (4/50) were infected compared to 6.4% (16/250) observed among blood donors (OR, 1.3; CI, 0.41 to 4.0;
P
= 0.43). Among subjects requiring bone marrow transplantation, seroprevalence rates were not elevated compared to those of blood donors (OR, 2.0; 95% CI, 0.10 to 122.9;
P
= 0.50). Because the need for high-quality KSHV detection methods are warranted and because questions remain about the optimal methods for assessing KSHV infection in individuals, we propose a systematic approach to standardize and optimize the assessment of KSHV infection rates using a combination of established and novel serological assays and methods.
Common Virulence Factors and Genetic Relationships between O18:K1:H7
Escherichia coli
Isolates of Human and Avian Origin
Authors: Moulin-Schouleur Maryvonne , Schouler Catherine , Tailliez Patrick , Kao Mu-Rong , Brée Annie , Germon Pierre , Oswald Eric , Mainil Jacques , Blanco Miguel , Blanco Jorge
Published: October 2006

Extraintestinal pathogenic (ExPEC)
Escherichia coli
strains of serotype O18:K1:H7 are mainly responsible for neonatal meningitis and sepsis in humans and belong to a limited number of closely related clones. The same serotype is also frequently isolated from the extraintestinal lesions of colibacillosis in poultry, but it is not well known to what extent human and avian strains of this particular serotype are related. Twenty-two ExPEC isolates of human origin and 33 isolates of avian origin were compared on the basis of their virulence determinants, lethality for chicks, pulsed-field gel electrophoresis (PFGE) patterns, and classification in the main phylogenetic groups. Both avian and human isolates were lethal for chicks and harbored similar virulence genotypes. A major virulence pattern, identified in 75% of the isolates, was characterized by the presence of F1 variant fimbriae; S fimbriae; IbeA; the aerobactin system; and genomic fragments A9, A12, D1, D7, D10, and D11 and by the absence of P fimbriae, F1C fimbriae, Afa adhesin, and CNF1. All but one of the avian and human isolates also belonged to major phylogenetic group B2. However, various subclonal populations could be distinguished by PFGE in relation to animal species and geographical origin. These results demonstrate that very closely related clones can be recovered from extraintestinal infections in humans and chickens and suggest that avian pathogenic
E. coli
isolates of serotype O18:K1:H7 are potential human pathogens.
Occurrence of Carbapenem-Resistant
Acinetobacter baumannii
Clones at Multiple Hospitals in London and Southeast England
Authors: Coelho Juliana M. , Turton Jane F. , Kaufmann Mary E. , Glover Judith , Woodford Neil , Warner Marina , Palepou Marie-France , Pike Rachel , Pitt Tyrone L. , Patel Bharat C. , Livermore David M.
Published: October 2006

From late 2003 to the end of 2005, the Health Protection Agency's national reference laboratories received approximately 1,600 referrals of
Acinetobacter
spp., including 419 and 58 examples, respectively, of two carbapenem-resistant
Acinetobacter baumannii
lineages, designated OXA-23 clones 1 and 2. Representatives of these clones were obtained from 40 and 8 hospitals, respectively, in London or elsewhere in Southeast England. Both clones had
bla
OXA-23
-like genes, as well as the intrinsic (but downregulated)
bla
OXA-51
-like carbapenemase genes typical of
A. baumannii
. Both were highly multiresistant: only colistin and tigecycline remained active versus OXA-23 clone 1 isolates; OXA-23 clone 2 isolates were also susceptible to amikacin and minocycline. These lineages increase the burden created by the southeast (SE) clone, a previously reported
A. baumannii
lineage with variable carbapenem resistance contingent on upregulation of the
bla
OXA-51
-like gene. Known since 2000, the SE clone had been referred from over 40 hospitals by the end of 2005, with 627 representatives received by the reference laboratories. The OXA-23 clone 2 is now in decline, but OXA-23 clone 1 continues to be referred from new sites, as does the SE clone. Their spread is forcing the use of unorthodox therapies, principally colistin and tigecycline, although the optimal regimens remain uncertain.
Expertise of Laboratories in Viral Load Quantification, Genotyping, and Precore Mutant Determination for Hepatitis B Virus in a Multicenter Study
Authors: Laperche Syria , Thibault Vincent , Bouchardeau Françoise , Alain Sophie , Castelain Sandrine , Gassin Michelle , Gueudin Marie , Halfon Philippe , Larrat Sylvie , Lunel Françoise , Martinot-Peignoux Michèle , Mercier Bernard , Pawlotsky Jean-Michel , Pozzetto Bruno , Roque-Afonso Anne-Marie , Roudot-Thoraval Françoise , Sauné Karine , Lefrère Jean-Jacques
Published: October 2006

A national evaluation study was performed in 14 specialized laboratories with the objective of assessing their capacities to provide (i) hepatitis B virus (HBV) viral loads (VL), (ii) HBV genotypes, and(iii) identification of precore/core mutants. The panel consisted of 12 HBV DNA-positive samples with VLs from 2.8 to 9.1 log10copies/ml, different HBV genotypes (A to F), and 3 mutant and 9 wild-type samples at nucleotide 1896. The coefficients of variation of the mean VLs ranged from 2.4% to 10.4% with the Cobas HBV Monitor assay, from 1.8% to 5.5% with the CobasTaqMan 48, from 1.5 to 26.2% with RealArt HBV PCR, and from 0 to 7% with branched DNA (bDNA). The Cobas Monitor assay underestimated the VLs of genotype F samples, with differences ranging from 1.4 to 2.4 log10copies/ml. The accuracies of genotype determinations ranged from 33% to 100%, and those of precore mutant determinations ranged from 25 to 100%. This study showed some drawbacks of two widely used assays: (i) Cobas Monitor has a narrow dynamic range and underestimates genotype F sample VLs and (ii) bDNA shows poor sensitivity and may fail to identify patients with low VLs. With higher performance in terms of analytical sensitivity combined with a larger dynamic range and an ability to quantify the main genotypes equally, real-time PCR methods appear more appropriate for accurate monitoring of HBV DNA quantification. Furthermore, the clinical implications of HBV genotyping and the determination of precore/core mutants need to be clearly stated to justify the standardization of these methods.
Genotyping of Measles Virus in Clinical Specimens on the Basis of Oligonucleotide Microarray Hybridization Patterns
Authors: Neverov Alexander A. , Riddell Michaela A. , Moss William J. , Volokhov Dmitriy V. , Rota Paul A. , Lowe Luis E. , Chibo Doris , Smit Sheilagh B. , Griffin Diane E. , Chumakov Konstantin M. , Chizhikov Vladimir E.
Published: October 2006

An oligonucleotide microarray hybridization method for identification of most known measles virus (MV) genotypes was developed. Like the conventional genotyping method, the microarray relied on detecting sequence differences in the 450-nucleotide region coding for the COOH-terminal 150 amino acids of the nucleoprotein (N). This region was amplified using PCR primers binding to all known MV genotypes. The microarray included 71 pairs of oligonucleotide probes (oligoprobes) immobilized on glass slides. Each pair consisted of a genotype-specific oligoprobe, which matched the sequence of only one target genotype, and a control oligoprobe, which contained mismatches at the nucleotide positions unique to this genotype. A pattern recognition algorithm based on cluster analysis of the ratios of hybridization signals from specific and control oligoprobes was used to identify the specific MV genotype. Following the initial validation, the method was used for rapid genotyping of two panels of coded samples. The results of this study showed good sensitivity (90.7%), specificity (100%), and genotype agreement (91.8%) for the new method compared to the results of genotyping conducted using phylogenetic analysis of viral sequences of the C terminus of the N gene. In addition, the microarray demonstrated the ability to identify potential new genotypes of MV based on the similarity of their hybridization patterns with those of known MV genotypes.
Genetic Background and Antibiotic Resistance ofStaphylococcus aureusStrains Isolated in the Republic of Georgia
Authors: Revazishvili Tamara , Bakanidze Lela , Gomelauri Tsaro , Zhgenti Ekaterine , Chanturia Gvantsa , Kekelidze Merab , Rajanna Chythanya , Kreger Arnold , Sulakvelidze Alexander
Published: October 2006

The genetic composition and antibiotic sensitivities of 50 clinical isolates ofStaphylococcus aureusobtained from various clinics in the Republic of Georgia were characterized.S. aureusstrains ATCC 700699 and ATCC 29737 were included as reference standards in all analyses. All 52 strains had identical 16S rRNA profiles. In contrast, pulsed-field gel electrophoresis (PFGE) identified 20 distinct PFGE types among the 52 strains examined, which indicates that PFGE is more discriminating than is 16S rRNA sequence analysis for differentiatingS. aureusstrains. The results of our PFGE typing also suggest that multiple genetic subpopulations (related at the ca. 85% similarity level, based on their SmaI PFGE patterns) exist among the GeorgianS. aureusstrains. Twenty-two of the 50 Georgian strains were methicillin resistant and PCR positive formecA, and 5 strains were methicillin sensitive even though they possessedmecA. None of the strains were vancomycin resistant or containedvanA. The nucleotide sequences ofmecAfragments obtained from allmecA-containing strains were identical. Our data indicate that the population ofS. aureusstrains in Georgia is fairly homogeneous and that the prevalence of methicillin-resistant,mecA-positive strains is relatively high in that country.
Age Dependence of Adenovirus-Specific Neutralizing Antibody Titers in Individuals from Sub-Saharan Africa
Authors: Thorner Anna R. , Vogels Ronald , Kaspers Jorn , Weverling Gerrit J. , Holterman Lennart , Lemckert Angelique A. C. , Dilraj Athmanundh , McNally Lisa M. , Jeena Prakash M. , Jepsen Soren , Abbink Peter , Nanda Anjali , Swanson Patricia E. , Bates Andrew T. , O'Brien Kara L. , Havenga Menzo J. E. , Goudsmit Jaap , Barouch Dan H.
Published: October 2006

We assessed neutralizing antibody titers to adenovirus serotype 5 (Ad5) and six rare adenovirus serotypes, serotypes 11, 35, 50, 26, 48, and 49, in pediatric populations in sub-Saharan Africa. We observed a clear age dependence of Ad5-specific neutralizing antibody titers. These data will help to guide the development of Ad vector-based vaccines for human immunodeficiency virus type 1 and other pathogens.
Comparative Study of Surgical Instruments from Sterile-Service Departments for Presence of Residual Gram-Negative Endotoxin and Proteinaceous Deposits
Authors: Lipscomb I. P. , Sihota A. K. , Keevil C. W.
Published: October 2006

The ineffective cleaning of surgical instruments may be a vector for the transmission of hospital-acquired infections. The aim of this research was to investigate whether further decontamination procedures need to be instigated in sterile-service departments (SSDs) to reduce the risk of nosocomial illnesses, such as endotoxemia, sepsis, or iatrogenic Creutzfeldt-Jakob disease (to date, 1,147 cases of confirmed Creutzfeldt-Jakob disease deaths in the United Kingdom since 1990 have been reported). Instrument sets were obtained from nine anonymous United Kingdom National Health Service (NHS) primary care trust SSDs. The investigation implemented an advanced light microscopy technique, episcopic differential interference contrast microscopy with the sensitive fluorescent reagents SYPRO Ruby and 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI), to detect proteinaceous and microbial contamination levels. Gram-negative lipopolysaccharide (LPS) endotoxin was monitored using a dansylated polymyxin B fluorochrome agent. None of the 260 instruments examined displayed signs of microbial colonization or LPS endotoxin contamination. However, over 60 percent of the instruments showed a high degree of protein soiling (0.4 to 4.2 μg protein/mm
2
). Some instruments appeared soiled with crystalline deposits that may consist of a potentially hazardous material contributing to inflammation and/or surgical shock. It is clear that the overall standard for cleaning must be raised in order to fulfill the imminent introduction of new European standards and to reduce the risk of cross-patient contamination and iatrogenic transmission.
Peritoneal mucinous cystadenocarcinoma of probable urachal origin: a challenging diagnosis
Authors: Gore D M , Bloch S , Waller W , Cohen P
Published: October 2006

This report describes the case of a mucinous cystadenocarcinoma of probable urachal origin that presented with mass effect, precipitating deep venous thrombosis and pulmonary embolism. The patient presented with acute symptoms of leg swelling, pain and dyspnoea, and a vague awareness of lower abdominal distension. Computer tomography showed a cystic mass closely related to the anterior abdominal wall and the superior aspect of the bladder. A 1500 cm3cyst adherent to the dome of the urinary bladder was resected on laparotomy. Partial cystectomy was not carried out in the belief that the cyst represented a benign lesion. Subsequent imaging has shown cystic changes in the anterior bladder wall, and the patient has been referred for partial cystectomy.
Patched homologue 1 mutations in four Japanese families with basal cell nevus syndrome
Authors: Matsuzawa N , Nagao T , Shimozato K , Niikawa N , Yoshiura K-i
Published: October 2006

Aim:To search for patched homologue 1 (PTCH1) mutations in four families with basal cell nevus syndrome (BCNS).

Methods:Mutation analysis ofPTCH1in unrelated Japanese families affected with BCNS was carried out by direct sequencing.

Results:Six novelPTCH1mutations, 833G→A in exon 6, 1415C→A and 1451G→T in exon 10, 2798delC in exon 17, 2918–2925dupAGTTCCCT in exon 18 and 3956C→A in exon 23, were identified.

Conclusions:Among the sixPTCH1mutations, two frameshift mutations (2798delC and 2918–2925dupAGTTCCCT) and one nonsense mutation (833G→A) are predicted to lead to premature termination ofPTCH1protein translation. Three simultaneous mutations, 1415C→A (A472D) and 1451G→T (G484V) in exon 10, and 3956G→A (R1319H) in exon 23, were found on one allele in only affected members in one family and none of them were found among 90 unrelated healthy Japanese. The three mutations on one chromosome may have resulted from errors in the recombinational repair process and this is the first report on thePTCH1mutations due to such a mechanism.
e19a2 BCR–ABL fusion transcript in typical chronic myeloid leukaemia: a report of two cases: Figure 1
Authors: Mondal B C , Majumdar S , Dasgupta U B , Chaudhuri U , Chakrabarti P , Bhattacharyya S
Published: October 2006

This report describes two patients with chronic myeloid leukaemia (CML): one of them developed accelerated phase CML and died 8 years after diagnosis and the other is at the chronic phase. Sequence analysis of reverse transcription-polymerase chain reaction products showed the presence of BCR–ABL fusion transcript e19a2. This finding suggests that CML carrying μ-BCR breakpoint may exhibit a clinical course similar to typical CML.
Splitting bone marrow trephines into frozen and fixed fragments allows parallel histological and molecular detection of B cell malignant infiltrates
Authors: Parrens M , Carrere N , Bouabdallah K , Fitoussi O , Goussot J-F , Dubus P , de Mascarel A , Merlio J-P
Published: October 2006

Clonality analysis of the immunoglobulin heavy chain (IgH) gene is helpful in identifying malignant B cell infiltrates in the bone marrow and is usually carried out on separate aspirates or on the same formalin-fixed decalcified bone marrow specimen. To determine whether the removal of the decalcification step would improve the molecular analysis, we first studied 12 bone marrow specimens with lymphoma infiltration split into a fixed and a small frozen fragment. Both the detection rate ofIgHgene monoclonality and DNA quality were found to be superior in the frozen part than in the fixed part. Conversely, to evaluate whether the split would compromise histological analysis, we selected a series of 134 bone marrow specimens obtained from patients with small B cell lymphoma and showing IgH monoclonality on the frozen part. The histological detection rate of infiltrated or suspicious infiltrates (95%) on the fixed part was not altered by saving a frozen part.
Expression of c-kit in human osteosarcoma and its relevance as a prognostic marker
Authors: Sulzbacher Irene , Birner Peter , Toma Cyril , Wick Nikolaus , Mazal Peter R
Published: October 2006

Aims:To examine the prognostic relevance of c-kit expression in human osteosarcomas and to evaluate the mutation status in exon 9 and exon 11 of the c-kit gene.

Methods:c-kit expression was examined in 100 human osteosarcomas by immunohistochemistry using paraffin embedded tumour tissues, and capillary sequencing of genomic DNA was performed to search for mutations in exons 9 and 11 of the c-kit gene.

Results:20 osteosarcomas showed c-kit expression ranging from 5% to 90% (mean 5.9%; SD 16.74%). Furthermore, DNA sequences of exon 9 and exon 11 of the c-kit gene were not altered in these tumours. Overall and disease free survival analysis did not reveal any differences between patients with osteosarcoma with c-kit expression and those with c-kit negative tumours.

Conclusions:C-kit expression is not a prognostic marker in patients with osteosarcoma. The protein expression is not linked to mutations in exon 9 or exon 11 of the c-kit gene. Therefore, these exons may not function as targets for treatment modalities based on the suppression of c-kit tyrosine kinase activity.
A laboratory strategy for genotyping haemoglobin H disease in the Chinese
Authors: Chan Amy Yuk-Yin , So Chi-Chiu , Ma Edmond Shiu-Kwan , Chan Li-Chong
Published: October 2006

Background:The thalassaemias are the commonest blood disorders worldwide, with South East Asia and southern China as areas of high prevalence. Accurate diagnosis of these disorders helps in clinical management with improved outcome.

Methods:The α-globin genotypes of 100 Chinese patients in Hong Kong with haemoglobin H (Hb H) disease were characterised. Single-tube multiplex gap-PCR was used to detect −−SEA, −α3.7and −α4.2, while Hb CS, Hb QS and codon 30 (ΔGAG) were identified by single-tube multiplex amplification refractory mutation system (ARMS). Automated direct nucleotide sequencing of the amplified α2- and α1-globin genes was performed to characterise other non-deletional α-thalassaemia determinants.

Results:In the 100 cases studied, 99 cases had −−SEAin combination with deletional α+-thalassaemia or non-deletional α-globin gene mutation involving the α2-globin gene. In 70 cases of the deletional form, 43 cases showed the genotype of (−−SEA/−α3.7) and 27 cases of (−−SEA/−α4.2). Three of the 27 cases of (−−SEA/−α4.2) were found to have Hb Q-Thailand linkedin-ciswith −α4.2. The remaining 30 cases were of non-deletional form with the following genotypes: 11 cases of (−−SEA/αHbCSα), 9 cases of (−−SEA/αHbQSα), 3 cases of (−−SEA/αcd30 (ΔGAG)α), 3 cases of (−−SEA/αcd31α), 2 cases of (−−SEA/αpoly-Aα), 1 case of (−−SEA/αHbWestmeadα) and 1 case of (−−non-SEA/αHbQSα).

Conclusions:Based on two rapid diagnostic tests, multiplex gap-PCR and multiplex ARMS, more than 90% of the cases were genetically characterised. This laboratory strategy should be widely applicable for genetic diagnosis of α-thalassaemia.
Hay fever and asthma symptoms in conventional and organic farmers in The Netherlands
Authors: Smit Lidwien A M , Zuurbier Moniek , Doekes Gert , Wouters Inge M , Heederik Dick , Douwes Jeroen
Published: October 2006

Background:Farming has been associated with respiratory symptoms and with protection against atopy. To date, effects of organic farming on respiratory health have not been studied.

Aims:To (1) compare hay fever and asthma-like symptoms in organic and conventional farmers and (2) assess associations between current and childhood farm exposures and respiratory health effects by conducting a survey.

Methods:Questionnaire data from 1205 conventional and 593 organic farmers were evaluated. Associations between health effects and farm exposures were assessed by logistic regression analyses.

Results:Organic farmers reported less wheezing with shortness of breath and slightly more hay fever than conventional farmers. However, organic farming was not an independent determinant of hay fever when adjusted for farming practices and potential confounders. Livestock farmers who grew up on a farm had a threefold lower prevalence of hay fever than crop farmers without a farm childhood (odds ratio (OR) 0.3, 95% confidence interval (CI) 0.1 to 0.5). Both crop farmers who grew up on a farm and livestock farmers who did not grow up on a farm had a reduced prevalence, although less pronounced and not statistically significant. Use of disinfectants containing quaternary ammonium compounds was positively related to hay fever (OR 2.1, 95% CI 1 to 4.4). No effects of farming practices were found for asthma.

Conclusions:Our study adds to the evidence that a farm childhood in combination with current livestock farming protects against allergic disorders. This effect was found for both organic and conventional farmers.
Randomized Study of Paclitaxel and Tamoxifen Deposition into Human Brain Tumors: Implications for the Treatment of Metastatic Brain Tumors
Authors: Fine Robert L. , Chen Johnson , Balmaceda Casilda , Bruce Jeffrey N. , Huang May , Desai Manisha , Sisti Michael B. , McKhann Guy M. , Goodman Robert R. , Bertino Joseph S. , Nafziger Anne N. , Fetell Michael R.
Published: October 2006

Purpose: Drug resistance in brain tumors is partially mediated by the blood-brain barrier of which a key component is P-glycoprotein, which is highly expressed in cerebral capillaries. Tamoxifen is a nontoxic inhibitor of P-glycoprotein. This trial assessed, in primary and metastatic brain tumors, the differential deposition of paclitaxel and whether tamoxifen could increase paclitaxel deposition.

Experimental Design: Patients for surgical resection of their primary or metastatic brain tumors were prospectively randomized to prior paclitaxel alone (175 mg/m2/i.v.) or tamoxifen for 5 days followed by paclitaxel. Central and peripheral tumor, surrounding normal brain and plasma, were analyzed for paclitaxel and tamoxifen.

Results: Twenty-seven patients completed the study. Based on a multivariate linear regression model, no significant differences in paclitaxel concentrations between the two study arms were found after adjusting for treatment group (tamoxifen versus control). However, in analysis for tumor type, metastatic brain tumors had higher paclitaxel concentrations in the tumor center (1.93-fold, P = 0.10) and in the tumor periphery (2.46-fold, P = 0.039) compared with primary brain tumors. Pharmacokinetic analyses showed comparable paclitaxel areas under the serum concentration between treatment arms.

Conclusions: Paclitaxel deposition was not increased with this tamoxifen schedule as the low plasma concentrations were likely secondary to concurrent use of P-450-inducing medications. However, the statistically higher paclitaxel deposition in the periphery of metastatic brain tumors provides functional evidence corroborating reports of decreased P-glycoprotein expression in metastatic versus primary brain tumors. This suggests that metastatic brain tumors may respond to paclitaxel if it has proven clinical efficacy for the primary tumor's histopathology.
Intralymphatic Dendritic Cell Vaccination Induces Tumor Antigen–Specific, Skin-Homing T Lymphocytes
Authors: Grover Amelia , Kim Grace J. , Lizée Gregory , Tschoi Mary , Wang Gang , Wunderlich John R. , Rosenberg Steven A. , Hwang Sam T. , Hwu Patrick
Published: October 2006

Purpose: The identification of tumor antigens recognized by cytotoxic and T helper lymphocytes has led to the development of specific cancer vaccines. Immunization with tumor antigen-pulsed dendritic cells has proved effective at eliciting elevated levels of tumor antigen–specific T cells in patient blood, but objective clinical responses remain rare, suggesting that vaccine-induced T cells are not trafficking optimally to site(s) of tumor burden. Accumulating evidence from animal models suggests that route of immunization can have a substantial influence on the subsequent migration of primed, activated T cells in vivo.

Experimental Design: In a clinical trial designed to elicit more effective cytotoxic T-cell mediated antitumor responses, metastatic melanoma patients were immunized directly via a peripheral intralymphatic route with autologous dendritic cells pulsed with HLA-A*0201-restricted melanoma-associated peptide antigens derived from MART-1 and gp100.

Results: Within 10 days of intralymphatic dendritic cell vaccination, four of six patients developed dramatic and diffuse erythematous rashes in sun-exposed areas of skin that showed extensive T-cell infiltration. CTLs grown from rash biopsies were strongly enriched for tumor antigen–specific T cells that had elevated expression of cutaneous lymphocyte antigen and chemokine receptor-6, consistent with a skin-homing phenotype. Of note, the only patient in the study with cutaneously localized disease showed a significant regression of metastatic lesions following the development of a surrounding rash.

Conclusions: The evidence presented here is consistent with immunization studies in animal models and supports the concept that T cells are “imprinted” in peripheral lymph node sites to express specific ligands and chemokine receptors that allow them to migrate to skin. Furthermore, the preferential migration of the T cells to sun-exposed cutaneous sites suggests that inflammation plays a critical role in this migration. These observations suggest that further study of the effects of immunization route and inflammation on T-cell migration in humans is warranted, and could lead to vaccination approaches that would more reliably direct trafficking of activated T cells to diverse sites of metastatic disease.
Most Early Disseminated Cancer Cells Detected in Bone Marrow of Breast Cancer Patients Have a Putative Breast Cancer Stem Cell Phenotype
Authors: Balic Marija , Lin Henry , Young Lillian , Hawes Debra , Giuliano Armando , McNamara George , Datar Ram H. , Cote Richard J.
Published: October 2006

Purpose: The presence of disseminated tumor cells (DTC) in the bone marrow of breast cancer patients is an acknowledged independent prognostic factor. The biological metastatic potential of these cells has not yet been shown. The presence of putative breast cancer stem cells is shown both in primary tumors and distant metastases. These cells with a CD44+CD24−/low phenotype represent a minor population in primary breast cancer and are associated with self-renewal and tumorigenic potential. Recognizing the potential effect of prevalence of putative stem cells among DTC, we evaluated the bone marrow DTC.

Experimental Design: We employed the double/triple-staining immunohistochemistry protocol and modified the established bone marrow cytokeratin (CK) staining protocol by adding steps for additional antigens, CD44 and/or CD24. We evaluated 50 bone marrow specimens, previously categorized as CK+ from early breast cancer patients. CK+ cells were examined for CD44 and CD24 expression by light microscopy, fluorescence microscopy, and spectral imaging.

Results: We detected the putative stem cell–like phenotype in all CK+ specimens. The mean prevalence of putative stem/progenitor cells was 72% and median prevalence was 65% (range, 33-100%) among the overall DTC per patient, compared with primary tumors where this phenotype is reported in <10% of cells.

Conclusions: This is the first evidence of the existence of the putative stem-like phenotype within the DTC in bone marrow in early breast cancer patients. All patients had a putative stem cell phenotype among the DTC and most individual DTC showed such phenotype. Future molecular characterization of these cells is warranted.
The GNAS1 T393C Polymorphism Is Associated with Disease Progression and Survival in Chronic Lymphocytic Leukemia
Authors: Frey Ulrich H. , Nückel Holger , Sellmann Ludger , Siemer Dörte , Küppers Ralf , Dürig Jan , Dührsen Ulrich , Siffert Winfried
Published: October 2006

Purpose: B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of monoclonal mature B cells. The G protein Gαs subunit has been linked to proapoptotic processes in cancer cell lines. The TT genotype of the GNAS1 T393C polymorphism is associated with increased Gαs transcript levels and a more favorable clinical course in different solid cancers.

Experimental Design: We retrospectively genotyped 144 patients with B-CLL to examine a potential association between T393C genotypes with progression-free survival (time from diagnosis to initiation of chemotherapy) and overall survival.

Results: The C-allele frequency in the patient group was 0.57 and not significantly different from that of healthy blood donors. Median progression-free survival was significantly different between genotypes (TT 130 months; TC 100 months; CC 31 months; P = 0.0066). Multivariable analysis showed that besides of ZAP-70 (P = 0.005) and Binet stage (P < 0.001), the T393C polymorphism was an independent prognostic factor for progression-free survival [hazard ratio (HR) CC versus TT 2.7; P = 0.010]. In Binet A stages, ZAP-70–positive patients with CC genotypes had a HR of 4.4 to receive first therapy compared with ZAP-70–negative patients with T-alleles (P = 0.0001). Regarding overall survival, CC genotypes (median overall survival, 197 months) were at highest risk for death compared with T-alleles (median overall survival, 310 months) in both univariate (HR, 4.8; P < 0.0001) and multivariable analysis (HR, 5.6; P = 0.002).

Conclusions: Here, we show that the GNAS1 T393C status is a novel independent prognostic marker in patients with B-CLL. These results could help to define patients who could benefit from an early individualized therapy.
Monitoring Tumor Glucose Utilization by Positron Emission Tomography for the Prediction of Treatment Response to Epidermal Growth Factor Receptor Kinase Inhibitors
Authors: Su Helen , Bodenstein Claudia , Dumont Rebecca A. , Seimbille Yann , Dubinett Steven , Phelps Michael E. , Herschman Harvey , Czernin Johannes , Weber Wolfgang
Published: October 2006

Purpose: The mechanisms underlying the sensitivity of non–small cell lung cancer to epidermal growth factor receptor (EGFR) kinase inhibitors are complex, and there are no established markers to accurately predict treatment outcome in individual patients.

Experimental Design: We investigated whether tumors responding to EGFR inhibitors can be identified by measuring treatment-induced changes in glucose utilization by positron emission tomography with the glucose analogue fluorodeoxyglucose (FDG-PET). We studied a panel of cell lines with a spectrum of sensitivity to EGFR kinase inhibitors. After incubation with the EGFR kinase inhibitor gefitinib for various time points, FDG uptake, glucose transport rates, and hexokinase activity were determined. FDG uptake in vivo was assessed by microPET imaging of tumor xenografts in mice.

Results: In gefitinib-sensitive cell lines, there was a dramatic decrease in FDG uptake as early as 2 hours after treatment. Immunoblots showed the translocation of glucose transporters (GLUT3) from the plasma membrane to the cytosol; glucose transport rates were reduced 2.6-fold at this time. There was also a modest reduction of hexokinase activity. These metabolic alterations preceded changes in cell cycle distribution, thymidine uptake, and apoptosis. MicroPET studies showed an up to 55% decrease of tumor FDG uptake in sensitive xenografts within 48 hours. In contrast, gefitinib-resistant cells exhibited no measurable changes in FDG uptake, either in cell culture or in vivo.

Conclusion: Glucose metabolic activity closely reflects response to gefitinib therapy. FDG-PET may be a valuable clinical predictor, early in the course of treatment, for therapeutic responses to EGFR kinase inhibitors.
Sensitization by Dietary Docosahexaenoic Acid of Rat Mammary Carcinoma to Anthracycline: A Role for Tumor Vascularization
Authors: Colas Séverine , Mahéo Karine , Denis Fabrice , Goupille Caroline , Hoinard Claude , Champeroux Pascal , Tranquart François , Bougnoux Philippe
Published: October 2006

Purpose: To investigate whether dietary docosahexaenoic acid (DHA), a peroxidizable polyunsaturated ω-3 fatty acids, sensitizes rat mammary tumors to anthracyclines and whether its action interferes with tumor vascularization, a critical determinant of tumor growth.

Experimental Design: Female Sprague-Dawley rats were initiated by N-methylnitrosourea to develop mammary tumors and then assigned to a control group (n = 18), receiving a supplementation of palm oil, or to a DHA group (n = 54), supplemented with a microalgae-produced oil (DHASCO, 1.5 g/d). The DHA group was equally subdivided into three subgroups with addition of different amounts of α-tocopherol. Epirubicin was injected weekly during 6 weeks after the largest tumor reached 1.5 cm2, and subsequent changes in the tumor surface were evaluated. Tumor vascularization was assessed by power Doppler sonography before and during chemotherapy.

Results: DHA and α-tocopherol were readily absorbed and incorporated into rat tissues. Epirubicin induced a 45% mammary tumor regression in the DHA-supplemented group, whereas no tumor regression was observed in the control group. In the DHA group, before chemotherapy was initiated, tumor vascular density was 43% lower than in the control group and remained lower during chemotherapy. Enhancement of epirubicin efficacy by DHA was abolished in a dose-dependent manner by α-tocopherol, and the same trend was observed for DHA-induced reduction in tumor vascular density.

Conclusions: Dietary DHA supplementation led to a reduction in tumor vascularization before the enhancement of any response to anthracyclines, suggesting that DHA chemosensitizes mammary tumors through an inhibition of the host vascular response to the tumor.
Peripheral Blood Stem Cell Tumor Cell Contamination and Survival of Neuroblastoma Patients
Authors: Corrias Maria Valeria , Haupt Riccardo , Carlini Barbara , Parodi Stefano , Rivabella Lucia , Garaventa Alberto , Pistoia Vito , Dallorso Sandro
Published: October 2006

Purpose: Contribution of peripheral blood stem cell (PBSC) contaminating tumor cells to subsequent relapse and overall survival of neuroblastoma patients remains controversial.

Experimental Design: Neuroblastoma cell contamination of 27 PBSC harvests from stage IV neuroblastoma patients was assessed by quantitative RT-PCR for both tyrosine hydroxylase (TH) and GD2 synthase (GD2-s). The effect of PBSC contamination on survival was then analyzed.

Results: Seven PBSC tested negative for both markers; 19 were positive for GD2-s, 6 for TH, with 5 positive for both. Survival of the 20 patients with positive PBSC did not differ from that of the patients with negative PBSC (log-rank test, P = 0.134 and 0.218 for event-free survival and overall survival, respectively). By considering the TH and GD2-s results independently, a borderline (P = 0.053) negative effect on event-free survival was observed in patients reinfused with GD2-s-positive PBSC. When the status at transplant was taken into account, only the event-free survival of the patients rescued when in complete remission with GD2-s-negative PBSC was better, although not significantly, than that of patients infused with GD2-s-positive PBSC.

Conclusions: Our results obtained in a small cohort of homogeneously treated stage IV patients suggest that patient survival is not affected by PBSC contamination with the exception of a borderline negative effect on event-free survival in patients rescued when in complete remission.
Inhibition of Tumor Growth with Antiangiogenic Cancer Vaccine Using Epitope Peptides Derived from Human Vascular Endothelial Growth Factor Receptor 1
Authors: Ishizaki Hidenobu , Tsunoda Takuya , Wada Satoshi , Yamauchi Mai , Shibuya Masabumi , Tahara Hideaki
Published: October 2006

Purpose: Antiangiogenic therapy is now considered to be one of promising approaches to treat various types of cancer. In this study, we examined the possibility of developing antiangiogenic cancer vaccine targeting vascular endothelial growth factor receptor 1 (VEGFR1) overexpressed on endothelial cells of newly formed vessels in the tumor.

Experimental Design: Epitope-candidate peptides were predicted from the amino acid sequence of VEGFR1 based on their theoretical binding affinities to the corresponding HLAs. The A2/Kb transgenic mice, which express the α1 and α2 domains of human HLA-A*0201, were immunized with the epitope candidates to examine their effects. We also examined whether these peptides could induce human CTLs specific to the target cells in vitro.

Results: The CTL responses in A2/Kb transgenic mice were induced with vaccination using identified epitope peptides restricted to HLA-A*0201. Peptide-specific CTL clones were also induced in vitro with these identified epitope peptides from peripheral blood mononuclear cells donated by healthy volunteers with HLA-A*0201. We established CTL clones in vitro from human peripheral blood mononuclear cells with HLA-A*2402 as well. These CTL clones were shown to have potent cytotoxicities in a HLA class I–restricted manner not only against peptide-pulsed target cells but also against target cells endogenously expressing VEGFR1. Furthermore, immunization of A2/Kb transgenic mice with identified epitope peptides restricted to HLA-A*0201 was associated with significant suppression of tumor-induced angiogenesis and tumor growth without showing apparent adverse effects.

Conclusions: These results strongly suggest that VEGFR1 is a promising target for antiangiogenic cancer vaccine and warrants further clinical development of this strategy.
The Combination of the Rexinoid, LG100268, and a Selective Estrogen Receptor Modulator, Either Arzoxifene or Acolbifene, Synergizes in the Prevention and Treatment of Mammary Tumors in an Estrogen Receptor–Negative Model of Breast Cancer
Authors: Liby Karen , Rendi Mara , Suh Nanjoo , Royce Darlene B. , Risingsong Renee , Williams Charlotte R. , Lamph William , Labrie Fernand , Krajewski Stan , Xu Xiaochun , Kim Heetae , Brown Powel , Sporn Michael B.
Published: October 2006

Purpose: We tested whether a selective estrogen receptor modulator (SERM) and a rexinoid are active for prevention and treatment in the mouse mammary tumor virus-neu mouse model of estrogen receptor–negative breast cancer.

Experimental Design: For prevention, mice were fed a powdered control diet, the SERM arzoxifene (Arz, 20 mg/kg diet), the rexinoid LG100268 (268, 30 mg/kg diet), or the combination for 60 weeks. In a second prevention study, mice were fed Arz (6 mg/kg diet), 268 (30 mg/kg diet), the combination of Arz and 268, the SERM acolbifene (Acol, 3 mg/kg diet), or the combination of Acol and 268 for 52 weeks. For the treatment studies, mice with tumors were fed combinations of a SERM and 268 for 4 weeks.

Results: The rexinoid 268 and the SERMs Arz and Acol, as individual drugs, delayed the development of estrogen receptor–negative tumors. Moreover, the combination of a SERM and 268 was strikingly synergistic, as no tumors developed in any mouse fed the combination of 268 and a SERM. Moreover, this drug combination also induced significant tumor regression when used therapeutically. These drugs did not inhibit transgene expression in vitro or in vivo, and the combination of Arz and 268 inhibited proliferation and induced apoptosis in the tumors.

Conclusion: The combination of a rexinoid and SERM should be considered for future clinical trials.
On “Field Significance” and the False Discovery Rate
Authors: Wilks D. S.
Published: October 2006

The conventional approach to evaluating the joint statistical significance of multiple hypothesis tests (i.e., “field,” or “global,” significance) in meteorology and climatology is to count the number of individual (or “local”) tests yielding nominally significant results and then to judge the unusualness of this integer value in the context of the distribution of such counts that would occur if all local null hypotheses were true. The sensitivity (i.e., statistical power) of this approach is potentially compromised both by the discrete nature of the test statistic and by the fact that the approach ignores the confidence with which locally significant tests reject their null hypotheses. An alternative global test statistic that has neither of these problems is the minimum p value among all of the local tests. Evaluation of field significance using the minimum local p value as the global test statistic, which is also known as the Walker test, has strong connections to the joint evaluation of multiple tests in a way that controls the “false discovery rate” (FDR, or the expected fraction of local null hypothesis rejections that are incorrect). In particular, using the minimum local p value to evaluate field significance at a level αglobal is nearly equivalent to the slightly more powerful global test based on the FDR criterion. An additional advantage shared by Walker’s test and the FDR approach is that both are robust to spatial dependence within the field of tests. The FDR method not only provides a more broadly applicable and generally more powerful field significance test than the conventional counting procedure but also allows better identification of locations with significant differences, because fewer than αglobal × 100% (on average) of apparently significant local tests will have resulted from local null hypotheses that are true.
Consolidation and Cure Simulations for Laminated Composites
Authors: Yan Xiangqiao
Published: October 2006

Advanced fiber reinforced polymer composites have been increasingly used in various structural components. One of the important processes to fabricate high performance laminated composites is an autoclave assisted prepreg lay-up. Since the quality of laminated composites is largely affected by the cure cycle, selection of the cure cycle for each application is important and must be optimized. Thus, some fundamental model of consolidation and cure processes is necessary to properly select suitable parameters for each application. This article develops a finite element code, which combines ‘the consolidation model’ with ‘the thermo-chemical model’ during the autoclave processing of composite materials. Numerical examples, including comparison of the present numerical results with one-dimensional (1-D) and 2-D analytical solutions, are given to indicate the accuracy and effectiveness of the finite element formulation. In addition, using the finite element code, the 1-D cure process of a laminate made of 52 plies of AS4/3501-6 unidirectional tape is simulated and some numerical results are compared with experimental results available in the literature.
Building a British Superman: Physical Culture in Interwar Britain
Authors: Zweiniger-Bargielowska Ina
Published: October 2006

The celebration of the 'body beautiful' in the 1920s and 1930s is commonly associated with fascist Italy or nazi Germany. Focusing on the physical culture movement, this article argues that the endeavour to build a 'superman' was not confined to fascist dictatorships or Britain's small fascist parties. The physical culture movement played an important role in cementing the link between manliness, physical fitness and patriotism in interwar Britain. Stimulated by the Edwardian 'national efficiency' campaign, physical culturalists represented physical fitness as an obligation of citizenship and a patriotic response to the needs of the British Empire. This aspiration acquired a renewed sense of urgency after the first world war. Despite a penchant for rituals and displays which some might read as fascist, the relationship between fascism and the physical culture movement was complex and contested. During the early 1930s, some physical culturalists championed fascist leaders and ideology and parts of the physical culture press moved to the extreme right. The effort to align British physical culturalists with fascism was unsuccessful, however, and in the late 1930s the physical culture press backed the centre-right National Government which launched a National Fitness Campaign in the face of a growing threat from nazi Germany.