22 research outputs found

    Cloning and characterization of an adenoviral vector for highly efficient and doxycycline – suppressible expression of bioactive human single – chain interleukin 12 in colon cancer

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    BACKGROUND: Interleukin-12 (IL-12) is well characterized to induce cellular antitumoral immunity by activation of NK-cells and T-lymphocytes. However, systemic administration of recombinant human IL-12 resulted in severe toxicity without perceptible therapeutic benefit. Even though intratumoral expression of IL-12 leads to tumor regression and long-term survival in a variety of animal models, clinical trials have not yet shown a significant therapeutic benefit. One major obstacle in the treatment with IL-12 is to overcome the relatively low expression of the therapeutic gene without compromising the safety of such an approach. Our objective was to generate an adenoviral vector system enabling the regulated expression of very high levels of bioactive, human IL-12. RESULTS: High gene expression was obtained utilizing the VP16 herpes simplex transactivator. Strong regulation of gene expression was realized by fusion of the VP16 to a tetracycline repressor with binding of the fusion protein to a flanking tetracycline operator and further enhanced by auto-regulated expression of its fusion gene within a bicistronic promoter construct. Infection of human colon cancer cells (HT29) at a multiplicity of infection (m.o.i.) of 10 resulted in the production of up to 8000 ng/10(6 )cells in 48 h, thus exceeding any published vector system so far. Doxycycline concentrations as low as 30 ng/ml resulted in up to 5000-fold suppression, enabling significant reduction of gene expression in a possible clinical setting. Bioactivity of the human single-chain IL-12 was similar to purified human heterodimeric IL-12. Frozen sections of human colon cancer showed high expression of the coxsackie adenovirus receptor with significant production of human single chain IL-12 in colon cancer biopsies after infection with 3*10(7 )p.f.u. Ad.3r-scIL12. Doxycycline mediated suppression of gene expression was up to 9000-fold in the infected colon cancer tissue. CONCLUSION: VP16 transactivator-mediated and doxycycline-regulated expression of the human interleukin-12 gene enables highly efficient and tightly controlled cytokine expression in human cancer. These data illustrate the potential of the described adenoviral vector system for the safe and superior expression of therapeutic genes in the treatment of colorectal cancer and other malignancies

    On the possibility of measuring relativistic gravitational effects with a LAGEOS-LAGEOS II-OPTIS-mission

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    In this paper we wish to preliminary investigate if it would be possible to use the orbital data from the proposed OPTIS mission together with those from the existing geodetic passive SLR LAGEOS and LAGEOS II satellites in order to perform precise measurements of some general relativistic gravitoelectromagnetic effects, with particular emphasis on the Lense-Thirring effect.Comment: Abridged version. 16 pages, no figures, 1 table. First results from the GGM01C Earth gravity model. GRACE data include

    Postinfectious T-lymphocytic enteral leiomyositis as a rare cause of chronic intestinal pseudoobstruction.

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    T-lymphocytic enteral leiomyositis (T-lel) is a rare disorder causing chronic intestinal pseudo-obstruction (CIPO), with cases predominantly being reported in the field of veterinary and pediatric medicine. Here, we present a case of T-lel-associated CIPO in an adult female, who initially presented with a paralytic ileus 2 weeks after a common gastroenteritis. The histological diagnosis was established through full-thickness bowel biopsy, exhibiting a dense lymphocytic infiltrate in the lamina muscularis of the intestinal wall. This case shows that T-lel can be a cause of chronic intestinal pseudo-obstruction not only in children but also in adults. A subsequent induction of an immunosuppressive therapy with steroids, azathioprine, and ultimately TNF-alpha-inhibiting antibodies led to a slow recovery and stable disease

    Evaluation of Use of Epstein-Barr Viral Load in Patients after Allogeneic Stem Cell Transplantation To Diagnose and Monitor Posttransplant Lymphoproliferative Disease

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    Epstein-Barr virus (EBV)-induced posttransplant lymphoproliferative disease (PTLD) continues to be a serious complication following transplantation. The aim of the present study was to evaluate the EBV load as a parameter for the prediction and monitoring of PTLD. The EBV load was analyzed by a quantitative competitive PCR with 417 whole-blood samples of 59 patients after allogeneic stem cell transplantation (SCT). The EBV load was positive for all 9 patients with PTLD and for 17 patients without PTLD. The viral loads of patients with manifest PTLD differed from the loads of those without PTLD (median loads, 1.4 × 10(6) versus 4 × 10(4) copies/μg of DNA; P < 0.0001). A threshold value of 10(5) copies/μg of DNA showed the best diagnostic efficacy (sensitivity, 87%; specificity, 91%). However, in patients with less than three major risk factors for PTLD, the positive predictive value of this threshold was rather low. One week prior to the manifestation of PTLD, the EBV load was as low in patients who developed PTLD as in patients without disease (median, 2.2 × 10(4) copies/μg of DNA; P was not significant). EBV DNA tested positive first at 20 to 71 days prior to the clinical manifestation of PTLD and occurred with the same delay after transplantation regardless of disease (median delay, 52 versus 63 days; P was not significant). EBV DNA was detected earlier in patients with primary infections than in those with reactivations (33 versus 79 days; P = 0.01), but the peak levels were similar in the two groups. EBV primary infection or EBV reactivation is frequent in patients after allogeneic SCT but results in PTLD only in a subgroup of patients. Although evaluation of the EBV load has limitations, the EBV load represents a valuable parameter to guide therapy
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