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Application of hMOF (Human Males Absent on the First) and small peptide of hMOF in preparing drug for alleviating or treating human arsenic poisoning

An arsenic poisoning and drug technology, applied in the field of biomedicine, can solve the problems of abnormal DNA repair reaction, chromosomal aberration, genome instability, etc.

Active Publication Date: 2015-07-29
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current findings show that removal of MOF in human cells not only causes a broad-based decrease in H4K16 acetylation, leading to abnormal DNA repair responses, but also leads to genomic instability, spontaneous chromosomal aberrations, cell cycle dysregulation, and nuclear Changes in morphology, etc. (Rea S, Xouri G, and Akhtar A. Males absent on the first (MOF): from flies to human. Oncogene, 2007, 26:5385-94.)

Method used

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  • Application of hMOF (Human Males Absent on the First) and small peptide of hMOF in preparing drug for alleviating or treating human arsenic poisoning
  • Application of hMOF (Human Males Absent on the First) and small peptide of hMOF in preparing drug for alleviating or treating human arsenic poisoning
  • Application of hMOF (Human Males Absent on the First) and small peptide of hMOF in preparing drug for alleviating or treating human arsenic poisoning

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Example 1: Detection of the influence of arsenic trioxide on hMOF and its enzyme activity

[0056] 【experiment procedure】:

[0057] (1) Prepare cells: inoculate (5×10 5 per well) 293T cells (human embryonic kidney cells, purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences) and inoculated (5×10 3 per well) HeLa cells (human cervical cancer cells, purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences), and a cell slide was added to each well. 2 mL of cell culture medium was added to each well of a 6-well plate, and 0.5 mL of cell culture medium was added to each well of a 24-well plate, and cultured in a cell culture incubator (Sanyo, Japan) until the cell coverage reached 30%.

[0058] (2) Drug treatment: 293T cells were cultured until the cell coverage rate reached 30%, and four wells were selected, and each well was replaced with a new 2mL cell culture medium premixed with different arsenic concentrations, so that the final con...

Embodiment 2

[0072] Example 2: Determination of arsenic compounds on hMOF gene expression, gene promoter region activation and hMOF protein expression at different time points

[0073] experiment procedure:

[0074] (1) RT-qPCR detection of the effect of arsenic compounds on hMOF gene expression

[0075] 293T cells (5×10 5 cells / well), after the cells were treated with 0.2-0.8 μM for 48 hours, the cell culture medium was discarded and washed once with PBS. Add 1 mL of RNAiso Plus to each well, pipette and mix with a pipette tip, incubate at room temperature for 5-10 min, add 200 μl of chloroform, shake vigorously for 15 seconds, and let stand at room temperature for 5 min. Then, the total RNA was sequentially extracted using chloroform-isopropanol-ethanol, and finally the RNA concentration and A260nm / 280nm light absorbance were measured by NanoDrop.

[0076] cDNA was obtained by reverse transcription using TAKARA kit PrimeScript Reverse Transcriptase (No.2680A), and then real-time quant...

Embodiment 3

[0090] Example 3: Direct binding of arsenic compounds to hMOF

[0091] experiment procedure:

[0092] (1) Preparation of arsenic-conjugated resin

[0093] After 50 μl of NHS-activated agarose was equilibrated with coupling buffer (0.1 M phosphate, 0.15 M sodium chloride), the coupling agent PAPAO (10 μmol) was added. After 4 hours on ice, it was further eluted with coupling buffer. The remaining epoxy active sites were blocked with 1 M ethanolamine for 20 min at room temperature. Finally, the agarose gel was eluted with coupling buffer again and stored at 4°C for future use (protected from light).

[0094] (2) Affinity purification of arsenic-conjugated resin

[0095] 20 μL of prepared arsenic-conjugated resin was reacted with 200 μL of whole cell extract at 4° C. for 12 hours. Eluted three times with PBS the next day. After centrifugation, 4xSDS loading buffer (Beijing Dingguo Biotechnology Co., Ltd. Cat No-WB-0081) was added. Proteins bound to the arsenic-conjugated r...

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Abstract

The invention relates to an application of hMOF and a small peptide of hMOF in preparing a drug for alleviating or treating human arsenic poisoning and belongs to the technical field of bio-pharmaceutics. HMOF protein and the regional small peptide (WLCEYCLKYMKYEKSYRFHLGQCQ) of the hMOF protein are determined to have an efficient and exclusive alleviation or depoisoning effect by using 293T (an immortalized human embryonic kidney epithelial cell strain transfected with an adenovirus E1A gene and simultaneously expressing an SV40 large T antigen) and a cell strain of HeLa (a cervical cancer cell) as research materials through intra-cellular transient transfection and means such as an immunoblotting experiment, a double fluorescent reporter gene testing experiment and an in-vitro acetylase activity testing experiment. The hMOF and the small peptide of the hMOF are further used for preparing the relevant depoisoning drug, the alleviating drug or a drug composition for treating the arsenic poisoning caused by arsenic acute and particularly chronic environmental pollution.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and specifically relates to the application of hMOF and its small peptides in the preparation of drugs for alleviating or treating human arsenic poisoning, and further for the preparation of related antidote drugs for the treatment of arsenic poisoning caused by acute arsenic, especially chronic environmental pollution , Relief medicament or pharmaceutical composition. Background technique [0002] As the main pollution source of drinking water, arsenic (also known as arsenic) has become a global environmental pollution problem. Nevertheless, arsenic compounds have been used clinically for centuries as medicines, especially as one of the most effective medicines for the treatment of acute promyelocytic leukemia. However, its application is greatly limited due to its toxic effects on the heart, liver, kidney, nervous system, etc. In addition, it has been found that long-term exposure or inta...

Claims

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Application Information

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IPC IPC(8): A61K38/45A61P39/02
Inventor 蔡勇金景姬刘达王飞武冬璐赵琳红
Owner JILIN UNIV
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