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Method for treating human malignant solid tumor by using hepatocyte nuclear factor-1alpha

A liver cell nuclear factor and solid tumor technology, applied in the field of solid tumor treatment, can solve the problems of unscreened differentiation therapy drugs or proteins, inability to differentiate tumors, and reduce tumorigenesis of cancer cells

Active Publication Date: 2012-05-30
上海赛迪夫医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, although it has been confirmed that some substances or genes can improve certain biological characteristics of tumor cells in vitro experiments (such as reduced proliferation and clone formation ability, up-regulated expression of some normal cell function genes), some substances have even been proved to reduce cancer cells. Tumor formation of cells in animals, but it is often found that these substances also affect normal cells (side effects), and cannot specifically induce differentiation of solid tumors in vivo
The present inventors have studied the regulatory effects of all-trans retinoic acid, somatostatin, tumor necrosis factor, and arsenic trioxide on the differentiation of liver cancer cell lines in vitro, but no drugs or proteins with definite therapeutic effects on differentiation have been screened out. These substances were also found to be ineffective in inducing differentiation of solid tumors

Method used

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  • Method for treating human malignant solid tumor by using hepatocyte nuclear factor-1alpha
  • Method for treating human malignant solid tumor by using hepatocyte nuclear factor-1alpha
  • Method for treating human malignant solid tumor by using hepatocyte nuclear factor-1alpha

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Experimental program
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Effect test

Embodiment 1

[0089] Detection of HNF1α gene expression in human liver tumor cell line by Realtime RT-PCR

[0090] 1. The commercially available conventional liver tumor cell lines Huh-7, Hep3B, MHCC-H, MHCC-L, PLC, YY, and 7721 were all mixed with 5×10 5 Inoculated in a six-well plate per plate, cultivated with fresh culture medium containing 10% fetal bovine serum, extracted cellular RNA the next day, and measured OD with a spectrophotometer 260 Values ​​were formulated into working concentrations (1 μg / μl and 0.1 μg / μl), and RNA integrity was detected by 1% agarose gel electrophoresis.

[0091] 2. Realtime RT-PCR: Take 4 μg RNA, 2 μl Random primer, and add DEPC water to 33 μl, set at 70°C for 5 minutes, and after 0°C for 5 minutes, add 10 μl 5×Buffer, 3 μl dNTP, 2 μl RNA reverse transcriptase and 2 μl RNase After mixing the inhibitors, place at 37°C for 1.5h to obtain the reverse transcription product. After diluting the reverse transcription product, take 1 μl as a template for Realti...

Embodiment 2

[0096] Realtime RT-PCR and immunohistochemical detection of HNF1α gene and protein expression in human liver cancer tissues and adjacent tissues

[0097] 1. Realtime RT-PCR: Trizol method to extract RNA from human liver cancer and paracancerous tissues, and measure its OD with a spectrophotometer 260 Values ​​were formulated into working concentrations (1 μg / μl and 0.1 μg / μl), and RNA integrity was detected by 1% agarose gel electrophoresis. 4 μg of RNA was taken for reverse transcription and Realtime PCR amplification (reverse transcription reaction, Realtime PCR reaction conditions and primer sequences were the same as above).

[0098] The results showed that: Among the 11 pairs of human liver cancer tissues / adjacent tissues, the expression of HNF1α in 7 cases (63.64%) of liver cancer tissues was lower than that of the adjacent tissues.

[0099] 2. Immunohistochemistry: Human liver cancer and paracancerous tissue wax block 4mm serial section, baked in 60 ℃ oven for 30min an...

Embodiment 3

[0102] Detection of HNF1α gene and protein expression in liver tissue of rat primary liver cancer model induced by DEN by immunohistochemical method

[0103] 1. DEN was injected intraperitoneally at 70 mg / kg to prepare rat primary liver cancer models. During the modeling process, the rats were sacrificed before modeling and at 10w, 18w, and 22w after modeling.

[0104] 2. Take rat liver tissue, fix it overnight in 10% neutral formalin, trim the tissue into 1.0×1.0×0.5cm block, soak in running water for 12h, ethanol gradient (50%-75%-80%-95% - absolute ethanol) dehydration, xylene transparent and dipped in wax to make tissue wax block. Immunohistochemical staining was performed after serial sectioning of wax blocks.

[0105] The results showed that the expression of HNF1α in rat liver tissue was gradually weakened with the prolongation of modeling time, and the expression of HNF1α in liver cancer tissue was the weakest.

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Abstract

The invention relates to a method for treating human malignant solid tumors by using hepatocyte nuclear factor-1alpha (HNF1alpha). Specifically, the invention relates to a method for inducing differentiation of human malignant solid tumor cells by using HNF1alpha, thereby treating malignant solid tumors. The study of the invention shows that tumor cells can be effectively induced to differentiate through regulation of the gene expression of malignant solid tumor cell HNF1alpha, thus providing a new means for induced differentiation therapy of tumors.

Description

technical field [0001] The invention relates to the fields of molecular biology, cell biology and medicine. Specifically, the present invention relates to the method and application of using Hepatocyte Nuclear Factor-1α (Hepatocyte Nuclear Factor-1α, HNF1α) to induce the differentiation of human malignant solid tumor cells, so as to apply it to the treatment of solid tumors. Background technique [0002] The treatment of malignant solid tumors is one of the current clinical difficulties, especially for malignant solid tumors that cannot be removed by surgery, there is still a lack of effective treatment methods in clinic. Selecting key proteins, molecules and genes closely related to the occurrence and development of tumor cells for specific targeted regulation is one of the core issues in the treatment of malignant solid tumors. [0003] Tumor differentiation therapy (differentiation therapy) is an important breakthrough in the clinical treatment of tumors in the past 20 y...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/17A61P35/00
CPCA61K48/00A61K38/17C07K14/47A61K38/1709A61P35/00
Inventor 谢渭芬曾欣林勇尹川陈岳祥
Owner 上海赛迪夫医药科技有限公司
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