Biomedical subjects

P J Selby

Publications and source records attributed to P J Selby.

At least 19 recordsLinked to original sources

PubMed2006

A novel mechanism of CD40-induced apoptosis of carcinoma cells involving TRAF3 and JNK/AP-1 activation.

Membrane-presented CD40 agonists can induce apoptosis in carcinoma, but not normal homologous epithelial cells, whereas soluble agonists are growth inhibitory but not proapoptotic unless protein synthesis is blocked. Here we demonstrate that membrane-presented CD40 ligand (CD154) (mCD40L), but not soluble agonists, triggers cell death in malignant human urothelial cells via a direct mechanism involving rapid upregulation of TNFR-associated factor (TRAF)3 protein, without concomitant upregulation of TRAF3 mRNA, followed by activation of the c-Jun N-terminal kinase (JNK)/activator protein-1 (AP-1) pathway and induction of the caspase-9/caspase-3-associated intrinsic apoptotic machinery. TRAF3 knockdown abrogated JNK/AP-1 activation and prevented CD40-mediated apoptosis, whereas restoration of CD40 expression in CD40-negative carcinoma cells restored apoptotic susceptibility via the TRAF3/AP-1-dependent mechanism. In normal human urothelial cells, mCD40L did not trigger apoptosis, but induced rapid downregulation of TRAF2 and 3, thereby paralleling the situation in B-lymphocytes. Thus, TRAF3 stabilization, JNK activation and caspase-9 induction define a novel pathway of CD40-mediated apoptosis in carcinoma cells.

Apoptosis↗

PubMed2006

Differential susceptibility to TRAIL of normal versus malignant human urothelial cells.

Comparing normal human urothelial (NHU) cells to a panel of six representative urothelial cell carcinoma (UCC)-derived cell lines, we showed that while TRAIL receptor expression patterns were similar, susceptibility to soluble recombinant crosslinked TRAIL fell into three categories. 4/6 carcinoma lines were sensitive, undergoing rapid and extensive death; NHU and 253J cells were partially resistant and HT1376 cells, like normal fibroblasts, were refractory. Both normal and malignant urothelial cells underwent apoptosis via the same caspase-8/9-mediated mechanism. Rapid receptor downregulation was a mechanism for evasion by some UCC cells. TRAIL resistance in malignant urothelial cells was partially dependent on FLIP(L) and was differentially mediated by p38(MAPK), whereas in normal cells, resistance was mediated by NF-kappaB. Importantly, extensive killing of UCC cells could be induced using noncrosslinked TRAIL after prolonged exposure, with no damage to their homologous, normal urothelial cell counterparts.

Apoptosis↗

PubMed2006

Rasch analysis of the dimensional structure of the Hospital Anxiety and Depression Scale.

The Hospital Anxiety and Depression Scale (HADS) has been used extensively in cancer patients to identify psychological distress. Reports of the factor structure and screening performance of the instrument vary. Rasch models allow an assessment of the structure of a questionnaire by identifying item fit. Removal of misfitting items may improve both the dimensionality and efficacy of screening questionnaires. A Rasch analysis of the HADS-T and subscales was used to explore the factor structure, dimensionality and screening efficacy. A total of 1855 patients completed a touchscreen version of the HADS, including 381 patients who had received a psychiatric interview (SCAN/PSE). These data were analysed using Rasch models, and the screening efficacy at identifying cases of psychological distress and anxiety and depression evaluated. The results demonstrated that the structure of the HADS-T and subscales was unidimensional. Three items from the HADS-T, and one from each of the subscales demonstrated misfit. Screening efficacy for the HADS-T and subscales was modest. However, removal of misfitting items had little impact on screening, demonstrating that items could potentially be omitted, if required. The item range covered a narrow spectrum of psychological distress, predominantly higher levels of distress. Additional items have to be added if screening for moderate to mild distress is to be improved for cancer patients.

Adult↗

PubMed2006

Novel urothelium specific gene expression identified by differential display reverse transcriptase-polymerase chain reaction.

PURPOSE: Understanding the molecular basis of differential gene expression among different tissues at various developmental stages and in neoplastic transformation is an important biological goal. The potential clinical applications of this improved understanding are more precise diagnosis of disease, prediction of prognosis, novel targeted therapies and prediction of response to therapy. MATERIALS AND METHODS: Differential display reverse transcriptase-polymerase chain reaction was used to compare gene expression in bovine urothelium to that in autologous lung, esophagus, liver and spleen. Products that appeared to have urothelial specific expression were sequenced and assessed for homology with known sequences. Ribonuclease protection assays were used to further confirm the expression pattern. RESULTS: A total of 32 discrete cDNAs were identified, including 3 products from genes known to be urothelium specific in their expression, 16 with significant homology to bovine, human or mouse expressed sequence tags and 5 with no sequence homology to any currently available sequence. Urothelium specific mRNA expression was confirmed for 3 genes by ribonuclease protection assays and one (Udd06) was further characterized as a urea transporter. CONCLUSIONS: The use of differential display reverse transcriptase-polymerase chain reaction and other complementary techniques for parallel gene expression analysis will permit the complete characterization of the urothelial transcriptome and help identify potential molecular targets for rationally targeted therapy.

Animals↗

PubMed2005

Transcriptional control of the human urothelial-specific gene, uroplakin Ia.

The transcriptional control elements of tissue-specific genes may be exploited in the design of therapeutic constructs for use in human gene therapy. The uroplakins are a family of four proteins which form the asymmetric unit membrane of the urothelium. We have cloned the human uroplakin Ia gene and defined its genomic structure and transcriptional start site. Using quantitative RT-PCR in an extended panel of normal tissues, we have demonstrated highly urothelial-specific expression of this gene. A Dual-Luciferase assay was used to assess the transcriptional activity of a variety of promoter fragments of the human uroplakin Ia gene. A highly specific promoter fragment (consisting of 2147 bp of 5'-flanking sequence, intron 1 and the 5' UTR) was identified which regulated urothelial-specific expression in vitro. The human uroplakin Ia promoter identified has potential use in future gene therapy strategies to restrict transgene expression to the urothelium.

5' Flanking Region↗

PubMed2005

Development and evaluation of an instrument to assess social difficulties in routine oncology practice.

PURPOSE: To develop and preliminarily evaluate a Social Difficulties Inventory (SDI) for use in oncology practice. METHODS: Item generation from patients (n = 96), staff (n = 49) and the literature. Questions constructed and pre-tested (n = 42) resulting in a 22-item questionnaire. Psychometric evaluation (n = 271) assessed frequency of endorsement, factor structure, summated scales and construct validity followed by randomisation to criterion validity or test-retest arms. RESULTS: Items met preset frequency of endorsement criteria. Factor analysis revealed a four-factor structure, three clearly definable, (1) Physical ability, (2) Providing for the family and (3) Contact with others, explaining 45.8% variance. Summated scales developed from these demonstrated good reliability (Cronbach's alpha > 0.7) and were used to test and confirmed construct validity. Several comparators from the Life Events and Difficulties Schedule (LEDS) and Cancer Rehabilitation Evaluation System Short Form (CARES-SF) tested criterion validity and majority agreements were moderate to good. Test-retest reliability was good with most kappa values > 0.6. One item was eliminated resulting in a 21-item questionnaire. CONCLUSION: The SDI is relevant and easy to understand. Initial psychometric evaluation was encouraging. Ongoing work to evaluate the clinical meaning and utility of the instrument and to examine the relationships between SDI scores and clinical outcomes will provide guidance about its usefulness as an assessment tool in routine oncology practice.

Factor Analysis, Statistical↗

PubMed2004

Maintenance treatment with interferon for advanced ovarian cancer: results of the Northern and Yorkshire gynaecology group randomised phase III study.

A randomised phase III trial was conducted to assess the role of interferon-alpha (INFalpha) 2a as maintenance therapy following surgery and/or chemotherapy in patients with epithelial ovarian carcinoma. Patients were randomised following initial surgery/chemotherapy to interferon-alpha 2a as 4.5 mega-units subcutaneously 3 days per week or to no further treatment. A total of 300 patients were randomised within the study between February 1990 and July 1997. No benefit for interferon maintenance was seen in terms of either overall or clinical event-free survival. We conclude that INF-alpha is not effective as a maintenance therapy in the management of women with ovarian cancer. The need for novel therapeutics or strategies to prevent the almost inevitable relapse of patients despite increasingly effective surgery and chemotherapy remains.

Adult↗

PubMed2003

Feasibility and compliance of automated measurement of quality of life in oncology practice.

PURPOSE: Systematic quality-of-life (QOL) assessment may have value in oncology practice by increasing awareness of a wide range of issues, possibly increasing detection of psychologic morbidity, social problems, and changes in physical status, and improving care and its outcomes. However, logistic problems are substantial. Automated systems solve many of these problems. We field-tested the feasibility and compliance that can be achieved using a computer touchscreen system in two consecutive studies. PATIENTS AND METHODS: In study 1, a prospective cohort of 272 patients was offered QOL assessment at each clinic appointment for 6 months. In study 2, all patients (N = 1,291) were offered QOL assessment as part of clinic routine during a 12-week period. RESULTS: In study 1, 82% of patients agreed to take part, but over time, compliance was poor (median, 40%; mean, 43%) and deteriorated with longer follow-up. In study 2, the overall compliance was greatly increased (median, 100%; mean, 70%), and compliance was retained over multiple visits. In study 1, compliance was better in younger patients, males, and socially advantaged patients, but was not affected by the presence of depression or anxiety, or QOL. In the second study, building on experience in the first study, data collection and storage in the computer system was excellent, achieving 98% of collected data stored in one center. In general, patients were comfortable with the computers and the approach. Data collection on the wards was more difficult and less complete than in clinics, especially for patients undergoing acute admissions. CONCLUSION: Feasibility with higher compliance was demonstrated in study 2, in which the data collection was integrated into routine care, and can be improved with further technical initiatives and education of staff.

Adult↗

PubMed2003

Adenovirus-mediated gene therapy for bladder cancer: efficient gene delivery to normal and malignant human urothelial cells in vitro and ex vivo.

Existing local therapies for superficial transitional cell carcinoma (TCC) of the bladder have limited success in preventing progression to life-threatening, muscle-invasive disease, and novel therapies are needed. Recent studies have raised doubts concerning the feasibility of adenovirus-mediated gene therapy for bladder cancer. We have therefore investigated adenoviral transduction of normal and malignant human urothelial cells, both as primary cultures and in intact epithelium. All 15 primary normal human urothelial cell lines tested were transduced in vitro by Adv-cmv-beta-gal at high efficiency, and better than most human TCC cell lines. Eight primary human TCC explants were also successfully transduced. In contrast, in intact normal urothelium, transduction efficiency was lower, and occurred only in superficial epithelial layers. Expression of the hCAR adenovirus receptor, however, occurred throughout the full thickness of urothelium. Transduction of human TCC biopsy specimens was at least as efficient as intact normal urothelium.We demonstrate for the first time that adenoviral transduction of both normal and malignant human urothelial cells is feasible. A physical barrier, rather than hCAR status, may be the main determinant of transduction of intact epithelium. Clinical trials of adenovirus-mediated gene therapy for superficial bladder cancer are warranted.

Adenoviruses, Human↗

PubMed2002

Social problems in oncology.

A study was undertaken to describe, evaluate and categorise the social problems experienced by cancer patients. Ninety-six adult cancer patients at all stages of disease participated in either a telephone focus group discussion, a face to face focus group or an individual interview which were tape recorded and transcribed. Six experts analysed the transcripts. A total of 32 social problems were identified categorized under eight headings plus four single items. The categories were: problems with (1) managing in the home, (2) health and welfare services, (3) finances, (4) employment, (5) legal matters, (6) relationships, (7) sexuality and body image and (8) recreation. Problems with relationships and communication were the most frequently reported with financial, employment, body image and domestic problems also being widely endorsed. Female groups, younger patient groups and groups where the aim of treatment was palliative reported more social problems than other groups. Social problems are common and important to cancer patients. The social problems identified in this study will contribute to an item pool generated for developing a Social Problems Inventory that may be included in patient centred assessment as part of routine oncology practice.

Adult↗

PubMed2002

Computer-based quality of life questionnaires may contribute to doctor-patient interactions in oncology.

It is well recognized that oncologists should consider patients' quality of life and functioning when planning and delivering anticancer treatment, but a comprehensive assessment of how a patient feels requires a thorough inquiry. A standardized measurement of patients' quality of life may support clinicians in identifying important problems for discussion during the limited time of the medical consultations. The aim of this study was to assess the feasibility of computer-administered individual quality of life measurements in oncology clinics with immediate feedback of results to clinicians and to examine the impact of the information on consultations. The study employed a prospective non-randomized design with pre-test post-test within subjects comparisons and involved three medical oncologists and 28 cancer patients receiving chemotherapy. The intervention consisted of completion of quality of life questionnaires before the consultations and informing clinicians of the results. The main outcome measures were patients' perceptions of the content of baseline and intervention consultations and satisfaction with communication. A qualitative analysis of clinicians' interviews was performed. When clinicians had the quality of life results they enquired more often about daily activities (Z= -2.71, P=0.007), emotional problems (Z= -2.11, P=0.035) and work related issues (Z= -1.89, P=0.058). There was an increase in the number of issues discussed during the intervention consultation (Z= -1.89, P=0.059). Patients were highly satisfied with both consultations. The computer measurement was well accepted by patients who felt that the questionnaires were a useful tool to tell the doctors about their problems. The clinicians perceived that the quality of life data broadened the range of the clinical inquiry and helped them identify issues for discussion. Having symptoms and functional problems expressed quantitatively on a scale was useful for detection of change over time.

Adult↗

PubMed2002

BCL10 in malignant lymphomas--an evaluation using fluorescence in situ hybridization.

BCL10 is a tumour suppressor gene originally cloned from a t(1;14)(p22;q32) breakpoint in a case of mucosa-associated lymphoid tissue (MALT) lymphoma. Translocations involving this gene, though uncommon, are sometimes encountered in MALT lymphomas. This gene is thought to play an important role in the development of malignant lymphomas. Fluorescence in situ hybridization (FISH) was therefore undertaken on 22 cases of malignant lymphoma of varying histology to establish the incidence of rearrangements involving the BCL10 gene. Initially, one case with a novel t(1;2)(p22;p12) translocation involving the BCL10 gene was identified, in a marginal zone lymphoma of the MALT type, and was reported elsewhere. Seven other cases were subsequently identified with abnormalities in the 1p region, including a translocation with a breakpoint in the 1p22 region in a case of lymphoblastic lymphoma. However, none of these involved the BCL10 gene. Mutation analysis of BCL10 was then performed on 57 cases of malignant lymphoma, including 17 MALT lymphomas, by single-strand conformational polymorphism (SSCP) analysis of tumour DNA. Tissue was obtained for mutation analysis for 12 of the 22 cases analysed by FISH. Selected cases with SSCP band shifts were further studied by direct sequencing. Polymorphisms were identified in eight cases, but no mutations of pathogenic significance were identified. Further RT-PCR and mutation analysis was performed on cDNAs from 12 cases (four MALT, seven diffuse large B-cell lymphoma, one Hodgkin's disease) in which DNA analysis had already been completed. This included the MALT lymphoma with the t(1;2)(p22;p12) rearrangement. Again, no mutations were identified in the coding sequence. This study confirms that rearrangements of the BCL10 gene are uncommon in lymphoma (1/22) and may be limited tothe MALT subtype of non-Hodgkin's lymphomas. It was also found that breakpoints or rearrangements in the 1p22 region do not necessarily involve the BCL10 gene. Moreover, the absence of mutations at both the DNA (0/60) and the mRNA (0/12) level indicates that this gene is not frequently inactivated by mutation, in those tumours in which it is not involved in translocations. Our findings suggest that the BCL10 gene is unlikely to have a frequent or key role in general lymphomagenesis.

Adaptor Proteins, Signal Transducing↗

PubMed2001

The response of human dendritic cells to recombinant adenovirus, recombinant Mycobacterium bovis Bacillus Calmette Guerin and biolistic methods of antigen delivery: different induction of contact-dependant and soluble signals.

Dendritic cells (DCs) are the most potent antigen presenting cells for inducing T-cell immune responses. The ability to grow human DCs from monocyte precursors provides an abundant source of these cells, which can be modified in vitro to present antigens. Re-administration of modified DCs to patients as vaccines has been shown in some cases to induce immune responses against cancer and infectious disease. Gene delivery to DCs provides an intracellular source of antigen for efficient and persistent loading of major histocompatibility complex (MHC) class I molecules. The aim of this study was to use monocyte-derived DCs (MD-DCs) from healthy donors to compare in vitro gene transfer, mediated by adenovirus, M. bovis Bacillus Calmette Guerin (BCG) and biolistic delivery. Efficiency of transfection and effect on DC phenotype, allostimulatory capacity and cytokine secretion was investigated. Adenovirus and BCG both showed a comparable ability to transfect MD-DCs, whereas the biolistic delivery by gene gun was unsuccessful in the reporter gene delivery. BCG transfection promoted MD-DC maturation as is apparent in the surface phenotype, allostimulatory capacity and cytokine secretion from cells. In comparison, adenovirus and biolistic delivery had a reduced effect on MD-DCs although enhancement of co-stimulatory and MHC molecule expression occurred in the cells of some donors. Both BCG and adenovirus represent useful vectors for gene transfer to human DCs. The effect of BCG on DC maturation may provide additional signals for the induction of antigen-specific T-cell responses.

Adenoviruses, Human↗

PubMed2001

Identification of genes involved in human urothelial cell-matrix interactions: implications for the progression pathways of malignant urothelium.

Interactions between epithelial cells and the extracellular matrix are central to tissue homeostasis and have a dynamic role in tissue remodeling and repair. Regulation of these pathways is balanced by positive and negative feedback elements, many of which have been implicated in the pathways of malignant progression. We have used differential display to identify genes that are up-regulated in normal human urothelial cells in response to exposure to extracellular matrix proteins (Matrigel) in vitro. This approach has identified genes that have key roles in cell-cell and cell-matrix interactions and that have been implicated in the progression of carcinomas of urothelial or other epithelial cell origins. One confirmed but unknown differentially expressed sequence was used to isolate a full-length gene, MIG-C4, from a human urothelial cDNA library. This gene was found to encode a novel urokinase plasminogen-activator receptor-like member of the Ly-6 family of glycosyl-phosphatidylinositol-anchored glycoproteins, and was identified as the human homologue of the rat metastasis-associated C4.4A gene. By in situ hybridization, MIG-C4 was expressed variably in normal urothelium and intensely in the tumor component of some noninvasive superficial lesions and in invasive and metastatic urothelial cancers. Thus, our approach has identified previously nonimplicated gene products involved in normal urothelium-matrix interactions that could be tumor-invasion or suppressor-gene targets in the development of invasive and metastatic tumor phenotypes.

Amino Acid Sequence↗

PubMed2001

Renal carcinoma cell lines inhibit natural killer activity via the CD94 receptor molecule.

MHC class I molecules protect normal and transformed cells from lysis by natural killer (NK) cells through recognition of receptors expressed on leucocytes. Defects in NK cell activity and lymphokine activated killer (LAK) cell generation have been previously demonstrated in patients with renal cell carcinoma (RCC). However, to date, the importance of NK receptor/MHC class I interactions for immune evasion by RCC cells has not been described. In this study, human RCC cell lines (HTB46, HTB47, ACHN, CRL 1933 and HTB44) were found to be susceptible to lysis by both NK cells and interleukin-15 (IL-15)-derived LAK cells from normal donors in vitro. However, when NK cells were co-cultured with RCC cells their expression of the CD94 NK receptor molecule was significantly increased and their cytolytic activity against RCC targets was reduced. The cytolytic activity of NK cells was restored by the addition of IL-15, which further augmented the expression of CD94 on CD56+ NK cells. Disruption of NK receptor-MHC class I interactions by the addition of blocking antibodies to CD94 had no effect on the lysis of K562 or HTB47 targets by NK cells. However, the sensitivity of HTB46 cells to NK-mediated lysis was increased by blocking the CD94 receptor molecule, but only when the NK cells had not been previously co-cultured with RCC cells. This was independent of the presence of IL-15. These results show that RCC cells can inhibit NK activity via CD94 and suggest that disruption of interactions between receptor and ligand on RCC cells in vivo may augment the immune response against tumours by innate effector cells.

Antibodies, Monoclonal↗

PubMed2001

The impact of attaining a minimal disease state after high-dose melphalan and autologous transplantation for multiple myeloma.

Initial studies with high-dose therapy (HDT) in myeloma suggest some beneficial effects of attaining a complete response (CR); however, the effect on survival is difficult to assess owing to inconsistencies in the definition of response between studies. We have analysed 96 newly diagnosed patients aged less than 65 years who received HDT and assessed the effect of response on survival using electrophoresis, immunofixation and fluorescent IgH polymerase chain reaction (PCR) to define CR. Patients received induction chemotherapy with C-VAMP (adriamycin, vincristine, methylprednisolone, cyclophosphamide) followed by melphalan 200 mg/m2 and reinfusion of peripheral blood stem cells. There was a high response to C-VAMP [CR = 24%, partial response (PR) = 64%], with all but one patient improving the depth of response after HDT (CR = 69%, PR = 31%). The progression-free survival (PFS) and overall survival (OS) were excellent at a median of 46.4 months and 72+ months. There was a trend towards an improved PFS in patients with an immunofixation-negative CR compared with patients with a PR (49.4 months, 41.14 months; P = 0.26). This was not evident when electrophoresis was used to define CR. The method used to define CR did not impact on the overall survival and fluorescent IgH PCR failed to add any additional prognostic information. This study supports the widespread use of the European Bone Marrow Transplantation group (EBMT) response criteria and suggests that immunofixation should be performed on all patients who become electrophoresis negative.

Adult↗

PubMed2001

Human dendritic cells genetically engineered to express a melanoma polyepitope DNA vaccine induce multiple cytotoxic T-cell responses.

PURPOSE: To assess the therapeutic potential of a melanoma polyepitope vaccine in human cells. Polyepitope DNA vaccines encoding T-cell epitopes have been demonstrated in murine systems to generate multiple cytotoxic T-cell responses to different antigens. Here, for the first time we demonstrate the ability of a melanoma polyepitope to stimulate lymphocytes from normal human donors to simultaneously generate multiple antigen-specific responses. EXPERIMENTAL DESIGN: Human dendritic cells (DC), transduced with a melanoma-polyepitope cDNA, were used to activate autologous lymphocytes from naïve donors as an in vitro model of DNA vaccination. Lymphocytes were primed with polyepitope or mock-transduced DC, boosted with peptide, then measured for antigen-specific cytotoxicity. RESULTS: Lymphocytes primed with polyepitope-transduced DC and boosted with peptide generated multiple cytotoxic responses. By contrast lymphocytes primed with mock-transfected DCs and boosted with peptide gave no specific cytotoxicity. However, when lymphocytes were repeatedly stimulated with polyepitope-transduced DCs immunodominance was seen with CTLs being generated to only one epitope, MART(27-35). CONCLUSIONS: We show in a human system that a melanoma polyepitope primes CTL to multiple epitopes. However, repeated stimulation by the polyepitope restricts the response to only the MART1 epitope. Thus, although polyepitope vaccines are an effective way of priming multiple naïve T-cell responses, continual boosting with polyepitope vaccines may, as a result of immunodominance, restrict the CTL. These findings have important implications for the use of DNA polyepitope vaccines in cancer immunotherapy.

Animals↗

PubMed2000

Proteomics: new perspectives, new biomedical opportunities.

Proteomics-based approaches, which examine the expressed proteins of a tissue or cell type, complement the genome initiatives and are increasingly being used to address biomedical questions. Proteins are the main functional output, and the genetic code cannot always indicate which proteins are expressed, in what quantity, and in what form. For example, post-translational modifications of proteins, such as phosphorylation or glycosylation, are very important in determining protein function. Similarly, the effects of environmental factors or multigenic processes such as ageing or disease cannot be assessed simply by examination of the genome alone. This review describes the underlying technology and illustrates several areas of biomedical research, ranging from pathogenesis of neurological disorders to drug and vaccine design, in which potential clinical applications are being explored.

Animals↗