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Biomedical subjects

D George

Publications and source records attributed to D George.

At least 19 recordsLinked to original sources
PubMed2001

Microarray analysis detects novel Pax3 downstream target genes.

Pax3 is a transcription factor that is required for the development of embryonic neural tube, neural crest, and somatic derivatives. Our previous study (Mayanil, C. S. K., George, D., Mania-Farnell, B., Bremer, C. L., McLone, D. G., and Bremer, E. G. (2000) J. Biol. Chem. 275, 23259-23266) reveals that overexpression of Pax3 in a human medulloblastoma cell line, DAOY, resulted in an up-regulation in alpha-2,8-polysialyltransferase (STX) gene expression and an increase in polysialic acid on neural cell adhesion molecule. This finding suggests that STX might be a previously undescribed downstream target of Pax3. Because Pax3 is important in diverse cellular functions during development, we are interested in the identification of additional downstream targets of Pax3. We utilized oligonucleotide arrays and RNA isolated from stable Pax3 transfectants to identify potential target genes. A total of 270 genes were altered in the Pax3 transfectants as compared with the vector control and parental cell line. An independent analysis by cDNA expression array and real-time quantitative polymerase chain reaction of several genes confirmed the changes observed by the oligonucleotide microarray data. Of the genes that displayed significant changes in expression, several contain paired and homeodomain binding motifs of Pax3 in their promoter regions. Using promoter-luciferase reporter transfection assays and electromobility shift assays, we showed at least one previously undescribed downstream target, STX, to be a biological downstream target of Pax3. Thus we report several previously undescribed candidate genes to be potential downstream targets of Pax3.

Animals↗
PubMed2001

Cytomegalovirus ventriculoencephalitis in a bone marrow transplant recipient receiving antiviral maintenance: clinical and molecular evidence of drug resistance.

We describe a case of CMV ventriculoencephalitis in a severely immunocompromised bone marrow transplant recipient who was receiving combination therapy with ganciclovir and foscarnet for treatment of viremia and retinitis. Analysis of sequential viral isolates recovered from the patient's cerebrospinal fluid suggested that disease developed because of the presence of viral resistance and, possibly, low tissue penetration of antiviral agents.

Antiviral Agents↗
PubMed2001

Paclitaxel, estramustine phosphate, and carboplatin in patients with advanced prostate cancer.

PURPOSE: To determine the safety and activity of weekly paclitaxel in combination with estramustine and carboplatin (TEC) in patients with advanced prostate cancer. PATIENTS AND METHODS: In a dose-escalation study, patients with advanced prostate cancer were administered paclitaxel (weekly 1-hour infusions of 60 to 100 mg/m(2)), oral estramustine (10 mg/kg), and carboplatin (area under the curve, 6 mg/mL-min every 4 weeks). Paclitaxel levels were determined 0, 30, 60, 90, and 120 minutes and 18 hours after infusion, and a concentration-time curve was estimated. Once a safe dose was established, a multi-institutional phase II trial was conducted in patients with progressive androgen-independent disease. RESULTS: Fifty-six patients with progressive androgen-independent disease were treated for a median of four cycles. The dose of paclitaxel was escalated from 60 to 100 mg/m(2) without the occurrence of DLT. Posttherapy decreases in serum prostate-specific antigen levels of 50%, 80%, and 90% were seen in 67%, 48%, and 39% (95% confidence interval, 55% to 79%, 35% to 61%, 26% to 52%) of the patients, respectively. Of the 33 patients with measurable disease, two (6%) had a complete response and 13 (39%) had a partial response. The overall median time to progression was 21 weeks, and the median survival time for all patients was 19.9 months. Major grade 3 or 4 adverse effects were thromboembolic disease (in 25% of patients), hyperglycemia (in 38%), and hypophosphatemia (in 42%). Significant leukopenia, thrombocytopenia, and peripheral neuropathy were not observed. CONCLUSION: TEC has significant antitumor activity and is well tolerated in patients with progressive androgen-independent prostate cancer.

Aged↗
PubMed2001

Quantitative analysis of transgene protein, mRNA, and vector DNA following injection of an adenoviral vector harboring glial cell line-derived neurotrophic factor into the primate caudate nucleus.

Gene therapy for neurodegenerative diseases relies on stable expression of a vector-mediated transgene in the human central nervous system (CNS). In nonhuman primate CNS, transgene expression has been primarily assessed using descriptive histological methods. Here, we quantified the expression of a human glial cell line-derived neurotrophic factor (hGDNF) transgene using an ELISA specific for hGDNF protein and real-time quantitative RT-PCR and PCR for hGDNF mRNA and vector DNA, respectively. Transgene expression was assessed 1 week after injection of an E1-, E3-deleted adenovirus harboring hGDNF into the caudate nucleus of St. Kitts green monkey. We found that 57-147 million and 116-771 million copies of hGDNF mRNA and vector DNA, respectively, were present per 10,000 copies of the beta-actin gene. In the same sites, 40-152 pg of hGDNF protein per milligram of tissue was measured. Comparisons of these measures among monkeys demonstrated variable vector DNA and protein levels, but consistent mRNA levels at one-third of the level of vector DNA. This suggests that local responses to the vector play a role in the level of transgene expression and that high levels of vector DNA do not necessarily predict a high level of transgene protein. However, the results of this study do show that neuroprotective levels of GDNF transgene expression can be achieved following injection of an adenoviral vector into nonhuman primate caudate. Moreover, these assays provide quantitative methods for evaluating and comparing viral vectors in primate CNS.

Actins↗
PubMed2001

Co-medication with hydrolytic enzymes in radiation therapy of uterine cervix: evidence of the reduction of acute side effects.

PURPOSE: The use of additional therapy with an oral enzyme preparation containing trypsin, chymotrypsin and papain has been suggested for the reduction of toxicity due to radiation therapy. This study was conducted to test the efficacy and tolerability of this enzyme combination in preventing or reducing the acute side effects of radiation therapy in patients with locally advanced cervical cancer. METHODS: A prospective, randomised, open, clinical trial was carried out on 120 patients (aged 24-85 years) with locally advanced, biopsy-proven carcinomas of the uterine cervix (stages IIa, lIb or IIIb). Patients received 50 Gy of external radiation therapy over a period of 5 weeks, followed by intracavitary brachytherapy (20-30 Gy). Patients assigned to the test group (60 patients) received additional treatment with enzymes. Patients were evaluated at weekly intervals for acute radiation therapy-related side effects, according to the RTOG/EORTC grading criteria, and then after the end of radiation therapy for another 8 weeks. Occurrence of adverse events, if any, was also recorded. RESULTS: The study revealed that the maximum extent of acute radiation side effects was reduced in the enzyme group: skin reactions (mean: 0.97 vs 1.68 in the control group, P < 0.001), vaginal mucosal reactions (0.55 vs 0.85, P = 0.10), genitourinary symptoms (0.93 vs 1.38, P < 0.001) and gastrointestinal reactions (1.12 vs 1.30, P = 0.12). The sum-scores during treatment, expressed as area under the curve, were significantly less in the enzyme treated patients. In the follow-up visits all observed side effects of radiation therapy were of lower intensity in the enzyme group than in the control group. CONCLUSIONS: In patients with locally advanced cancer of the uterine cervix, oral enzyme therapy was found to be effective in significantly reducing radiation therapy-related side effects such as genitourinary symptoms, subcutaneous changes and reactions of the vaginal mucosa.

Acute Disease↗
PubMed2001

Ganglioside modulates ligand binding to the epidermal growth factor receptor.

Whereas previous investigations have shown that pharmacologic addition of gangliosides inhibits keratinocyte proliferation by downregulating epidermal growth factor receptor phosphorylation, the underlying biochemical basis and physiologic relevance are unknown. Using Scatchard and displacement plots, we have shown that supplemental purified gangliosides decrease the binding of (125)I-labeled epidermal growth factor to keratinocyte-derived SCC12 cells. Conversely, SCC12 cells transfected with sialidase and thus depleted of gangliosides show increased ligand binding to the epidermal growth factor receptor, which is consistent with their increased proliferation in response to epidermal growth factor and transforming growth factor-alpha, and increased phosphorylation of the epidermal growth factor receptor, and downstream signal transduction pathway components. The mechanism of the altered binding appears to involve primarily decreased numbers of available receptors within the intact membrane, but not altered receptor protein expression. These studies provide evidence that the effect of gangliosides on keratinocyte proliferation results, at least in part, from the direct binding of ganglioside to the receptor and disruption of the receptor-ligand interaction. Manipulation of membrane ganglioside content may be a powerful new means to alter epidermal growth factor receptor-dependent cell proliferation.

DNA, Complementary↗
PubMed2001

Platelet-derived growth factor receptors: a therapeutic target in solid tumors.

Platelet-derived growth factor (PDGF) was one of the first polypeptide growth factors identified that signals through a cell surface tyrosine kinase receptor (PDGF-R) to stimulate various cellular functions including growth, proliferation, and differentiation. Since then, several related genes have been identified constituting a family of ligands (primarily PDGF A and B) and their cognate receptors (PDGF-R alpha and beta). To date, PDGF expression has been shown in a number of different solid tumors, from glioblastomas to prostate carcinomas. In these various tumor types, the biologic role of PDGF signaling can vary from autocrine stimulation of cancer cell growth to more subtle paracrine interactions involving adjacent stroma and even angiogenesis. The tyrosine kinase inhibitor imatinib mesylate (formerly STI571, [Gleevec]; Novartis Pharmaceuticals Corp, East Hanover, NJ) blocks activity of the Bcr-Abl oncoprotein, and was recently approved for several indications in the treatment of chronic myeloid leukemia. Imatinib mesylate is also a potent inhibitor of the PDGF-R kinase and is currently being evaluated for the treatment of PDGF-responsive tumors such as prostate cancer. More clinical trials that investigate both established clinical endpoints of response and benefit, as well as surrogate endpoints that may describe the biologic significance of PDGF-R inhibition in vivo are needed to expand the applications that target the PDGF axis.

Antineoplastic Agents↗
PubMed2000

Heightened CD40 ligand gene expression in peripheral CD4+ T cells from patients with kidney allograft rejection.

BACKGROUND: CD40 ligand (CD40L) gene expression is increased in rejecting allograft biopsies. We, therefore, tested the possibility that CD40L gene expression is heightened in peripheral CD4+ T cells during renal allograft rejection. METHODS: CD40L gene expression in peripheral blood CD4+ T cells from two renal transplant groups was measured by quantitative reverse transcription-polymerase chain reaction (RT-PCR). Group 1: 20 patients with excellent renal transplant function; group 2: 25 patients with findings of acute and/or chronic allograft nephropathy (CAN); and group 3 of 12 normal controls. CD4+ cells were isolated by positive selection. The modifying effect of cyclosporine (CsA) and FK506 on CD40L gene expression was further tested in vitro in CD4+ T lymphocytes separated from pokeweed mitogen- (PWM) activated peripheral blood lymphocytes (PBL) preparations. RESULTS: Mean+/-SD expressions of CD40L gene, in aM, in groups 1-3 were as follows: 0.0052+/-0.0094; 0.022+/-0.023 (P=0.0038 vs. group 2); and 0.014+/-0.005. Levels of CD40L gene expression correlated significantly with acute rejection Banff 97 score (R2=0.44, P=0.0004) and severity of intertubular capillary changes (ITCC) (R2=0.33, P=0.011). After in vitro activation, CD40L gene expression increased by approximately 4-fold and the addition of CsA or FK506 diminished CD40L gene expression to a base level. CONCLUSION: Peripheral CD4+ T cell CD40L gene expression increases significantly in acute rejection and CAN and may serve as a non-invasive method to monitor allograft function and determine the biological response to CsA and FK506.

Acute Disease↗
PubMed2000

Overexpression of murine Pax3 increases NCAM polysialylation in a human medulloblastoma cell line.

Polysialic acid (PSA) is a developmentally regulated carbohydrate found primarily on neural cell adhesion molecules (NCAM) in embryonic tissues. The majority of NCAM in adult tissues lacks this unique carbohydrate, but polysialylated NCAM (PSA-NCAM) is present in adult brain regions where neural regeneration persists and in some pediatric brain tumors such as medulloblastoma, which show greater propensity for leptomeningeal spread. Pax3, a developmentally regulated paired homeodomain transcription factor, is thought to be involved in the regulation of neural cell adhesion molecules. Overexpression of murine Pax3 into a human medulloblastoma cell line (DAOY) resulted in an increase in NCAM polysialylation and a 2-4-fold increase in alpha2, 8-polysialyltransferase type II mRNA levels. No difference was observed in alpha2,8-polysialyltransferase type IV message. The addition of PSA to NCAM changed the adhesive behavior of these Pax3 transfectants. Transfectants expressing high PSA-NCAM show much less NCAM-dependent aggregation than those with less PSA-NCAM. In addition, Pax3 transfectants having high PSA-NCAM show heterophilic adhesion involving polysialic acid to heparan sulfate proteoglycan and agrin. These observations suggest that a developmentally regulated transcription factor, Pax3, could affect NCAM polysialylation and subsequently cell-cell and cell-substratum interaction.

Animals↗
PubMed2000

Modulation of EGF receptor activity by changes in the GM3 content in a human epidermoid carcinoma cell line, A431.

Gangliosides have been described as modulators of growth factor receptors. For example, GM3 addition in cell culture medium inhibits epidermal growth factor (EGF)-stimulated receptor autophosphorylation. Furthermore, depletion of ganglioside by sialidase gene transfection appeared to increase EGF receptor (EGFR) autophosphorylation. These data suggested that changes in GM3 content may result in different responses to EGF. In this study, the ceramide analog d-threo-1-phenyl-2-decannoylamino-3-morpholino-1-propanol ([D]-PDMP), which inhibits UDP-glucose-ceramide glucosyltransferase, and addition of GM3 to the culture medium were used to study the effects of GM3 on the EGFR. Addition of 10 microM [D]-PDMP to A431 cells resulted in significant GM3 depletion. Additionally, EGFR autophosphorylation was increased after EGF stimulation. When exogenous GM3 was added in combination with [D]-PDMP, the enhanced EGFR autophosphorylation was returned to control levels. [D]-PDMP also increased EGF-induced cell proliferation, consistent with its effect on autophosphorylation. Once again, the addition of GM3 in combination with [D]-PDMP reversed these effects. These results indicate that growth factor receptor functions can be modulated by the level of ganglioside expression in cell lines. Addition of GM3 inhibits EGFR activity and decrease of GM3 levels using [D]-PDMP treatment enhances EGFR activity. Modulation of growth factor receptor function may provide an explanation for how transformation-dependent ganglioside changes contribute to the transformed phenotype.

Carcinoma, Squamous Cell↗
PubMed2000

Gene expression analysis by massively parallel signature sequencing (MPSS) on microbead arrays.

We describe a novel sequencing approach that combines non-gel-based signature sequencing with in vitro cloning of millions of templates on separate 5 microm diameter microbeads. After constructing a microbead library of DNA templates by in vitro cloning, we assembled a planar array of a million template-containing microbeads in a flow cell at a density greater than 3x10(6) microbeads/cm2. Sequences of the free ends of the cloned templates on each microbead were then simultaneously analyzed using a fluorescence-based signature sequencing method that does not require DNA fragment separation. Signature sequences of 16-20 bases were obtained by repeated cycles of enzymatic cleavage with a type IIs restriction endonuclease, adaptor ligation, and sequence interrogation by encoded hybridization probes. The approach was validated by sequencing over 269,000 signatures from two cDNA libraries constructed from a fully sequenced strain of Saccharomyces cerevisiae, and by measuring gene expression levels in the human cell line THP-1. The approach provides an unprecedented depth of analysis permitting application of powerful statistical techniques for discovery of functional relationships among genes, whether known or unknown beforehand, or whether expressed at high or very low levels.

Cell Line↗
PubMed2000

Stem cell transplantation for the treatment of Fanconi anaemia using a fludarabine-based cytoreductive regimen and T-cell-depleted related HLA-mismatched peripheral blood stem cell grafts.

We have employed a new cytoreductive regimen to transplant two patients with Fanconi anaemia (FA), using T cell-depleted two HLA-allele disparate related peripheral blood stem cell transplants (PBSCTs). Patient 1, a 5-year-old male with FA and aplastic anaemia, initially received an HLA two-antigen mismatched unrelated cord blood transplant and failed to engraft. He received fludarabine (Flu) and cyclophosphamide (Cy), followed by a CD34(+) E-rosette(-) (CD34(+)E(-)), T cell-depleted, granulocyte colony-stimulating factor (G-CSF)-mobilized PBSCT from his HLA B-DRB1 mismatched father. He received anti-thymocyte globulin (ATG), steroids, FK506 and G-CSF after transplant for rejection and graft-versus-host disease (GVHD) prophylaxis. The patient is now 23 months after SCT with no evidence of GVHD and with full haematopoietic and immune reconstitution. Patient 2, a 10-year-old boy with FA and myelodysplastic syndrome, received single-dose total body irradiation (SDTBI), Flu and Cy followed by a CD34(+)E(-), T-cell-depleted, G-CSF-mobilized PBSCT from his HLA B-DRB1 mismatched sister. He also received ATG, steroids, FK506 and G-CSF after transplant. The patient is now 12 months after SCT in complete remission with no evidence of GVHD. Absolute neutrophil counts (ANC) of > 1 x 10(9)/l were achieved on day 11 and day 10 post transplant respectively. Both patients are fully engrafted. In summary, we report two successful T-cell-depleted stem cell transplants from mismatched related donors for the treatment of Fanconi anaemia, using a fludarabine-based cytoreduction. Both patients experienced minimal toxicity, rapid engraftment and no GVHD.

Child, Preschool↗
PubMed2000

Diagnostic value of cervical discography in the management of cervical discogenic pain.

We did a retrospective analysis of patients who had a total of 46 cervical discs examined by cervical discography and then evaluated the postoperative results of 14 of the 16 patients who underwent cervical spine fusion at those levels in accordance with the results of our discograms. The average symptomatic period prior to discography was 12 months, and cervical discogenic pain was successfully localized in all 16 patients. Immediately post-cervical discectomy and anterior fusion, subjectively, all 14 patients had good to excellent results, and after a mean follow-up of 6.5 (1.5 to 14) months, 11 patients (78.6%) continued with good to excellent results and three patients (21.4%) developed related pain patterns. Thus, cervical discography, in a "select group" of patients with chronic intractable neck pain but negative or indeterminate imaging findings who are being considered for surgical intervention, can help localize the symptomatic level and potentially benefit the patients by surgical intervention.

Adult↗
PubMed1999

Severity of depression in abstinent alcoholics is associated with monoamine metabolites and dehydroepiandrosterone-sulfate concentrations.

Depressed mood increases the relapse risk of abstinent alcoholics; its neurobiological correlates may include reduced serotonin and norepinephrine turnover rates and increased cortisol concentrations during detoxification stress. Neurosteroids such as dehydroepiandrosterone and its sulfate (DHEA and DHEA-S) may antagonize cortisol action and may have mood-elevating effects on their own. We measured severity of depression with Beck's Depression Inventory (BDI) and Hamilton's Depression Rating Scale (HDRS), plasma concentrations of cortisol, DHEA and DHEA-S, and CSF concentrations of the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA), the norepinephrine metabolite 3-methoxy-4-hydroxyphenylglycol (MHPG) and the dopamine metabolite homovanillic acid (HVA) in 21 abstinent alcoholics after 4 weeks of abstinence and in 11 age-matched healthy control subjects. Only CSF MHPG concentrations were reduced in alcoholics compared to control subjects (41.4 +/- 6.6 vs. 53.3 +/- 8.6 pmol/ml). Self-rated depression was significantly correlated with CSF MHPG (Spearman's R = +0.57, P < 0.01), CSF 5-HIAA (R = +0.51, P < 0.05) and plasma cortisol concentrations (R = +0.50, P < 0.05). Negative correlations were found between DHEA-S concentrations and both self-rated depression (R = -0.45, P < 0.05) and observer-rated depression (R = -0.55, P < 0.05). The ratio of DHEA-S to cortisol serum concentrations was also negatively correlated with depression (BDI: R = -0.55, P < 0.01; HDRS: R = -0.63, P < 0.005). Anxiety (Spielberger's State Anxiety Scale) was only associated with CSF MHPG concentrations (R = +0.58, P < 0.01). Our findings point to the importance of noradrenergic dysfunction in the pathogenesis of depression among abstinent alcoholics and indicate that their mood states may also be modulated by a low DHEA-S to cortisol ratio, hypothetically indicative of low stress protection capacities.

Adult↗
PubMed1999

Comparison of immune reconstitution after unrelated and related T-cell-depleted bone marrow transplantation: effect of patient age and donor leukocyte infusions.

Unrelated bone marrow transplantation (BMT) is often complicated by fatal opportunistic infections. To evaluate features unique to immune reconstitution after unrelated BMT, the lymphoid phenotype, in vitro function, and life-threatening opportunistic infections after unrelated and related T-cell-depleted (TCD) BMT were analyzed longitudinally and compared. The effects of posttransplant donor leukocyte infusions to treat or prevent cytomegalovirus (CMV) or Epstein-Barr virus (EBV) infections on immune reconstitution were also analyzed. This study demonstrates that adult recipients of TCD unrelated BMTs experience prolonged and profound deficiencies of CD3(+), CD4(+), and CD8(+) T-cell populations when compared with pediatric recipients of unrelated BMT and adults after related BMT (P <.01), that these adults have a significantly increased risk of life-threatening opportunistic infections, and that the rate of recovery of CD4 T cells correlates with the risk of developing these infections. Recovery of normal numbers of CD3(+), CD8(+), and CD4(+) T-cell populations is similar in children after related or unrelated BMT. This study also demonstrates that adoptive immunotherapy with small numbers of unirradiated donor leukocytes can be associated with rapid restoration of CD3(+), CD4(+), and CD8(+) T-cell numbers, antigen-specific T-cell responses, and resolution of CMV- and EBV-associated disease after unrelated TCD BMT.

Adolescent↗
PubMed1999

Sialidase gene transfection enhances epidermal growth factor receptor activity in an epidermoid carcinoma cell line, A431.

Glycosphingolipids expressed in cancer cells have been implicated in the modulation of tumor cell growth through their interaction with transmembrane signaling molecules such as growth factor receptors. For glycosphingolipids to interact with growth factor receptors, the presence of sialic acid seems to be essential. Stable transfection of a gene encoding a soluble Mr 42,000 sialidase into a human epidermoid carcinoma cell line (A431) provided an approach by which the level of terminal lipid-bound sialic acid on the cell surface could be altered. In the sialidase-positive clones, the level of ganglioside GM3 was diminished, and little change was observed in protein sialylation. Sialidase-transfected cells grew faster than control cells. Sialidase expression did not modify the binding of epidermal growth factor (EGF) to its receptor but enhanced EGF receptor (EGFR) tyrosine autophosphorylation as compared to that of parental cells or cells transfected with the vector (pcDNA3) alone. Moreover, the phosphorylation of the EGFR, as well as other protein substrates, was observed at low EGF concentrations, suggesting an increase in the receptor kinase sensitivity. These data provided evidence that changes in ganglioside expression in cancer cells by appropriate gene transfection can dramatically affect EGFR kinase activity. Hence, the modulation of ganglioside expression may represent an approach to alter tumor cell growth.

Carcinoma, Squamous Cell↗