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ABSTRACT: Antibody-based fusion proteins are the next generation of antibody therapies for cancer and other diseases. CD20 antigen, which is overexpressed on cell membranes in nearly 95% of cases of B-cell Non-Hodgkin's Lymphoma, is an attractive target for the therapy of B-lymphoid malignancies. Lidamycin (LDM) is a potent enediyne-containing antitumor antibiotic that now has entered phase II clinical trials. In this study, we prepared an engineered fusion protein, scFv-LDP, consisting of an anti-CD20 scFv fragment and the apoprotein LDP of LDM using DNA recombination. After purification and refolding, scFv-LDP was found to bind specifically to CD20-positive lymphoma cells using ELISA and indirect immunofluorescent cytochemical staining assays. The energized fusion protein scFv-LDP-AE was obtained using molecular reconstitution of the active chromophore AE of LDM and scFv-LDP. MTT assay revealed potent cytotoxicity of scFv-LDP-AE to CD20-positive Raji and Daudi cells, with IC(50) values of 1.21×10(-11) and 6.24×10(-11) mol L(-1), respectively. An in vivo subcutaneous xenograft model of CD20-positive B cell lymphoma in BALB/c (nu/nu) mice was also utilized. Drugs were given intravenously on day 14 and 21 after tumor transplantation. In terms of maximal tolerated doses, scFv-LDP-AE at 0.3 mg kg(-1) suppressed tumor growth by 79.3%, and LDM at 0.05 mg kg(-1) by 68.6% (P<0.05). Results suggested scFv-LDP-AE could be a potential candidate for tumor-targeting therapy.
Science China. Life sciences 03/2011; 54(3):255-62. · 2.02 Impact Factor
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ABSTRACT: This study is to investigate the inhibitory effect of lidamycin (LDM) and its combination with methotrexate (MTX) on lung metastasis of fibrosarcoma by bioluminescence imaging in athymic mice. A stable luciferase transfected HT-1080 cell line was constructed and the capability to establish experimental lung metastasis in athymic mice was confirmed. The optical imaging system was applied to evaluate the formation of lung metastasis in vivo. In addition, metastatic nodules were counted for the evaluation of inhibition rates. As shown, the fluorescent intensity of luciferase-transfected HT-1080 cells was colinear with the cell population and the minimal detected cell population was 100 cells/well. Optical imaging showed that the fluorescent intensity of treated group was apparently lower than that of the control. The inhibition rates of lung metastasis by LDM alone at 0.025 mg x kg(-1) and 0.05 mg x kg(-1) were 53.9% and 75.9%, respectively, while that of MTX alone at 0.5 mg x kg(-1) was 70.2%. The combination of LDM at 0.025 mg x kg(-1) and MTX at 0.5 mg x kg(-1) showed an inhibition rate of 88.7%. The coefficient of drug interaction (CDI) was 0.82. The results herein demonstrated that LDM alone had strong anti-metastasis effect on human fibrosarcoma HT-1080 and the inhibition efficacy is strengthened when combined with MTX.
Yao xue xue bao = Acta pharmaceutica Sinica 01/2011; 46(1):45-9.
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ABSTRACT: To conduct an in vivo optical imaging analysis of the biodistribution of antibody Rituximab in lymphoma tumor-bearing nude mice.
Laser scanning confocal microscope and flow cytometry were employed to determine the affinity of FITC-labeled Rituximab (FITC-Rituximab) with human lymphoma Raji cells. And the in vivo optical imaging system was used to analyze the biodistribution of FITC-Rituximab in lymphoma-transplanted xenograft nude mice.
The results of flow cytometry and laser scanning confocal microscope demonstrated that FITC-Rituximab had remarkable affinity with lymphoma Raji cells and was mainly bound at cell membrane. The results of in vivo imaging analysis suggested that FITC-Rituximab could specifically accumulated at peritumor tissue less than 1 h, then penetrated into the interior of tumor and concentrated in 3-4 h. And the specific concentration of FITC-Rituximab could still been observed more than 8-10 h whereas there was no apparent fluorescence at other tissues. Furthermore, the results observed from a two-flank tumor xenograft model showed that FITC-Rituximab possessed specific binding affinity for CD20-overexpressed lymphoma.
The in vivo optical imaging system can accurately monitor the distribution of FITC-Rituximab in tumor-bearing nude mice. And this technique has a reference value and significance for a real-time analysis of tumor-targeting capability of antibody drugs.
Zhonghua yi xue za zhi 09/2010; 90(33):2367-70.
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ABSTRACT: To investigate the role of the extracellular domain of human 4-1BBL (ex4-1BBL) in regulating the in vitro activities of peripheral blood lymphocytes (PBL).
Viable cells were quantified with Trypan-blue exclusion assay. CytoTox 96 Nonradioactive Cytotoxicity Assay Kit was used for measuring LDH levels of supernatant. ELISA kit was used for measuring IL-2 level. In vitro cytotoxity of PBL combined with anti-CD3/anti-Pgp bispecific diabody plus ex4-1BBL was analyzed with CytoTox 96 nonradioactive method.
Ex4-1BBL can increase the proliferation of PBL, reduce cell death, promote IL-2 secretion, and the experimental group with ex4-1BBL showed obviously enhanced cytotoxic effect toward K562/A02 cells.
Ex4-1BBL may be an important adjuvant for improving activities of PBL.
Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology 06/2008; 24(5):431-3.
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Hongxing Guo,
Wenguo Jiang,
Wenge Liu,
Yingdai Gao,
Ming Yang,
Yuan Zhou,
Jinhong Wang,
Jing Qi, Xin Cheng,
Zhenping Zhu,
Chunzheng Yang,
Dongsheng Xiong
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ABSTRACT: Our previous data have shown a significantly higher tumor response to anti-CD3/anti-Pgp bispecific diabody-mediated immunotherapy for P-glycoprotein (Pgp)-overexpressing K562/A02 cells, but a rapid tumor relapse occurred at 1 week after therapy. In an attempt to overcome tumor recurrence, we supplemented the previous therapy with extracellular domain of human 4-1BBL (ex4-1BBL) to regulate the activation of peripheral blood lymphocyte (PBL). As a result, this combination showed enhanced cytotoxicity in vitro and eradicated the multidrug-resistant xenografts of K562/A02 in nude mice. Furthermore, no tumor recurrence was observed within 100 days after the first treatment. Therefore, when used as an adjuvant, ex4-1BBL may improve the outcome of PBL-based immunotherapy.
Cellular Immunology 03/2008; 251(2):102-8. · 1.97 Impact Factor
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Hongxing Guo,
Wenguo Jiang,
Wenge Liu,
Yingdai Gao,
Ming Yang,
Yuan Zhou,
Jinhong Wang,
Jing Qi, Xin Cheng,
Zhenping Zhu,
Chunzheng Yang,
Dongsheng Xiong
[show abstract]
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ABSTRACT: Our previous data have shown a significantly higher tumor response to anti-CD3/anti-Pgp bispecific diabody-mediated immunotherapy for P-glycoprotein (Pgp)-overexpressing K562/A02 cells, but a rapid tumor relapse occurred at 1 week after therapy. In an attempt to overcome tumor recurrence, we supplemented the previous therapy with extracellular domain of human 4-1BBL (ex4-1BBL) to regulate the activation of peripheral blood lymphocyte (PBL). As a result, this combination showed enhanced cytotoxicity in vitro and eradicated the multidrug-resistant xenografts of K562/A02 in nude mice. Furthermore, no tumor recurrence was observed within 100 days after the first treatment. Therefore, when used as an adjuvant, ex4-1BBL may improve the outcome of PBL-based immunotherapy.
Cellular Immunology.
