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Application of Kir2.1 as target spot to preparation of reagent or medicine for treating, preventing or diagnosing medulloblastoma

A medulloblastoma and drug technology, applied in anti-tumor drugs, drug combinations, biological tests, etc., can solve problems such as growth retardation, endocrine dysfunction, cognitive function decline, etc.

Pending Publication Date: 2022-01-04
THE FIRST AFFILIATED HOSPITAL OF ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, due to the side effects of radiotherapy, survivors of medulloblastoma often have cognitive decline, mental decline, growth retardation, endocrine dysfunction, infertility, and second tumor stimulation (Sun Xiaofei, Donnie Yen. Children's medulloblastoma Expert consensus on multidisciplinary diagnosis and treatment of tumors (CCCG-MB-2017)[J]. Chinese Journal of Pediatric Hematology and Tumors, 2018, 23(4))

Method used

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  • Application of Kir2.1 as target spot to preparation of reagent or medicine for treating, preventing or diagnosing medulloblastoma
  • Application of Kir2.1 as target spot to preparation of reagent or medicine for treating, preventing or diagnosing medulloblastoma
  • Application of Kir2.1 as target spot to preparation of reagent or medicine for treating, preventing or diagnosing medulloblastoma

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Through immunohistochemistry and data analysis, the results were as follows figure 1 shown.

[0024] Immunohistochemical staining

[0025] 1. Slicing: After all paraffin specimens are uniformly numbered, put them in a -20°C refrigerator for pre-freezing, and then perform the steps of slicing, spreading and air-drying. The thickness of the slices was all 3 μm.

[0026] 2. Dewaxing: Place the paraffin sections on an iron shelf, bake in an oven at 60°C for 45 minutes, and then quickly soak them in xylene for 15 minutes. Put it in another vat of xylene and soak for 15 minutes. Then soak in 100% alcohol for 10 minutes, 95% alcohol for 5 minutes, 85% alcohol for 5 minutes, 75% alcohol for 5 minutes, tap water for 5 minutes, and PBS for 5 minutes.

[0027]3. Antigen retrieval: Rinse with PBS 3 times, 5 min each time. Put the slices in a pressure cooker filled with restoration solution, heat to 100°C, cover the lid, and apply high pressure for 2min30s. Then rinse the cool...

Embodiment 2

[0057] Invasion migration and Western Blot experiments were carried out, and the results were as follows figure 2 It can be seen.

[0058] Medulloblast culture

[0059] The medulloblasts used in this experiment were all cultured in DMED medium containing 10% FBS, and kept in the cell incubator CO 2 Conditions of content 5%, temperature 37°C, humidity 95%. When the cell growth confluence reached 80%-90%, it was digested and subcultured with trypsin cell digestion solution, and both MB428 and MB913 cells were subcultured in 1 passage and 3 passages. Half of each cell was cryopreserved when it was transferred to the second passage, and the cells from the third to fifth passage were used for experiments.

[0060] Establishment of a medulloblast cell line stably silencing Kir2.1

[0061] 1. Kir2.1-shRNA interference lentivirus and nonsense sequence control lentivirus provided by Santa Cruze Company.

[0062] 2. For medulloblasts in the logarithmic growth phase, after washing ...

Embodiment 3

[0114] Co-immunoprecipitation experiments were carried out, and the results were as follows image 3 shown.

[0115] co-immunoprecipitation

[0116] The co-immunoprecipitation experiment used the Co-IP kit of Thermo Company, and the specific operation steps were as follows:

[0117] 1. Antibody and resin cross-linking

[0118] (1) Mix the cross-linked resin gently, pipette 80 μL into the chromatography column, centrifuge at 1000×g for 1 min, and wash twice with Couplingbuffer.

[0119] (2) Mix 10 μg of primary antibody or corresponding IgG, 10 μL of sodium cyanoborohydride and an appropriate volume of Coupling buffer to form 200 μL of antibody cross-linking mixture, seal the bottom of the chromatography column, and add the mixture to the chromatography column, 4 Rotational crosslinking at ℃ for 2-4h:

[0120] 2. Cell collection, lysis and pre-clearing

[0121] (1) When the medulla overexpressing Kir2.1 cells are cultured until the confluence reaches 80%, the medium is rem...

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Abstract

The invention belongs to the technical field of biology, and particularly relates to application of Kir2.1 as a target spot to preparation of a reagent or a medicine for treating, preventing or diagnosing medulloblastoma. The invention has important clinical application value for diagnosis, treatment and prevention of WNT / SHH medulloblastoma.

Description

technical field [0001] The invention belongs to the field of biotechnology, and specifically relates to the application of Kir2.1 as a target in the preparation of reagents or medicines for treating, preventing or diagnosing medulloblastoma. Background technique [0002] Medulloblastoma (MB) is the most common embryonic malignant brain tumor of the central nervous system in childhood, accounting for about 25% of all intracranial tumors in children. Currently, the treatment strategy for medulloblastoma is surgery combined with whole brain and spinal cord radiotherapy and adjuvant chemotherapy after radiotherapy. Stratified treatment is carried out according to risk factors. At this stage, the five-year recurrence-free survival rate of standard-risk medulloblastoma is about 70%-80%, and the five-year recurrence-free survival rate of high-risk medulloblastoma is about 60% (Sun Xiaofei , Zhen Zijun. Expert consensus on multidisciplinary diagnosis and treatment of medulloblastom...

Claims

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Application Information

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IPC IPC(8): G01N33/574G01N33/68A61K45/00A61P35/00
CPCG01N33/57407G01N33/57484G01N33/68A61K45/00A61P35/00
Inventor 蓝茜王艳霞党微旗缪静雅周红向东方崔有宏卞修武
Owner THE FIRST AFFILIATED HOSPITAL OF ARMY MEDICAL UNIV
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