Pentamidine and death domain receptor ligand united application

A combined application, pentamidine technology, applied in the field of medicine, can solve the problem of enhancing the sensitivity of leukemia cells to induce apoptosis, and achieve the effect of treating leukemia and enhancing sensitivity

Inactive Publication Date: 2010-08-25
SHANTOU UNIV MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the research on pentamidine enhancing the sensitivity of leukemia cells to death domain ligand-induced apoptosis has not been reported or patent application

Method used

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  • Pentamidine and death domain receptor ligand united application
  • Pentamidine and death domain receptor ligand united application
  • Pentamidine and death domain receptor ligand united application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The preparation and cell culture of embodiment 1 pentamidine and TRAIL

[0022] Stock solutions of 1 mg / ml pentamidine from Sigma and 1 mg / ml TRAIL from PeproTech EC were prepared in ddH2O. K562 adopts conventional cell culture, RPMI-1640 culture solution containing 10% calf serum, 100u / ml of penicillin and streptomycin, at 37°C, saturated humidity, 5% CO 2 Cultured in an incubator, and the medium was changed every 3 to 4 days. Collect logarithmic growth phase cells, adjust the number of cells to 1×10 5 / ml, take 10mL to the culture dish, add pentamidine with the final concentration of 0.5, 1, 2, 5, 10 μg / ml respectively to act on K562 cells for 48 or 72 hours. The rest of the conditions are exactly the same. Incubate at 37°C and collect cells.

Embodiment 2

[0023] Embodiment 2 Pentamidine inhibits cell proliferation activity

[0024] Tetramethylazolium salt (MTT) was purchased from Sigma. Prepare a 5 mg / ml stock solution with deionized water and store it at -20°C for later use. Take 100ul of each group of cells described in Example 1 and place them in a 96-well ELISA plate, then add 10ul of MTT storage solution to each well, incubate in a cell culture incubator at 37°C for 4 hours, take it out, add 100ul of isopropanol containing HCI, and mix The optical density (OD) was measured with a microplate reader with a wavelength of 570nM. The cell proliferation inhibition rate was calculated by (control OD-treatment OD) / control OD×%. Pentamidine has a certain inhibitory effect on cell proliferation at a concentration above 2ug / ml. For this reason, the range of 2-10 μg / ml pentamidine was used in combination with TRAIL.

Embodiment 3

[0025] Example 3 Pentamidine Enhances Sensitivity of Cells to TRAIL and Induces Apoptosis

[0026] The cells were pretreated with 10 μg / ml pentamidine for 20 hours, followed by 200 ng / ml TRAIL for 4 hours. In addition, corresponding controls were set up, such as no drug, 10 μg / ml pentamidine for 24 hours, and 200 ng / ml TRAIL for 4 hours. Cell morphological changes were observed with a light microscope. There was no significant change in cells when pentamidine or TRAIL was used alone, but the morphology of cells appeared apoptosis characteristics after pretreatment with pentamidine.

[0027] Preliminarily use 5, 10 μg / ml pentamidine to act on the cells for 20 hours, and then use 200ng / ml TRAIL for 4 hours, and set up corresponding controls. If no drug is added, 5, 10ug / ml pentamidine for 24 hours, 200ng / ml TRAIL for 4 hours The number of apoptotic cells was determined by Annexin V FITC / PI double-labeled apoptotic cells with flow cytometry (FACS) using the Annexin V apoptosis k...

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Abstract

The present invention belongs to the pharmaceutical field. The present invention particularly relates to the medicine of pentamidine combined with the legend of the death structure receptor in treating the tumor and the leukemia. The ligand of the death structure receptor in the present invention comprises TRAIL, TNF-alpha or the analogues, which is the combination of the pentamidine with the metabolite or the analogues. The benefit of the present invention is that the pentamidine sensibility of the leukemia cell of the ligand of the death structure receptor. In the clinical field, when thepentamidine of the therapeutic drug concentration is applied in the leukemia cell, which is tolerant to the ligand of the death structure receptor, the apoptosis can be induced; therefore the purposefor the leukemia treatment is realized.

Description

technical field [0001] The invention belongs to the field of drugs, in particular to a drug for treating leukemia with pentamidine combined with a death domain receptor ligand. technical background: [0002] Leukemia is a malignant tumor of the hematopoietic system. Since the first leukemia patient achieved clinical remission with chemotherapy in 1947, the treatment of leukemia has developed from simple chemotherapy to the comprehensive application of hematopoietic stem cell transplantation, biological therapy (including cytokines, immunotherapy, etc.), but so far The effect of leukemia treatment is still unsatisfactory. Therefore, how to establish new low-toxicity and high-efficiency leukemia treatment-related strategies is still the focus of researchers. In the normal human hematopoietic system, cell growth maintains a steady state, determined by the balance between cell proliferation and renewal and cell death. When this homeostasis imbalance is disrupted, leukemia and...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K31/155A61K45/06A61P35/00
Inventor 刘小珊蒋纪恺
Owner SHANTOU UNIV MEDICAL COLLEGE
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