Pharmaceutical composition for removing residual liver cancer stem cells with combined use of arsenic trioxide and all-transretinoic acid and application thereof

An all-trans retinoic acid, liver cancer stem cell technology, applied in drug combinations, active ingredients of hydroxyl compounds, anti-tumor drugs, etc., can solve the problems of unsatisfactory anti-cancer effect, unclear treatment mechanism of HCC, and stagnation of anti-HCC research. Achieve the effect of removing liver cancer stem cells and reducing dosage

Inactive Publication Date: 2019-02-01
金世龙
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In 2004, the State Food and Drug Administration (SFDA) approved As 2 o 3 Can be used to treat advanced HCC, however, multi-center phase II clinical trials in the treatment of advanced HCC showed that As alone 2 o 3 The anticancer effect is very unsatisfactory, and As 2 o 3 The mechanism of treatment of HCC is unclear, after which As 2 o 3 Anti-HCC research is almost at a standstill
ATRA treatment of APL targets the RARa part of the PML-RARa fusion oncoprotein, and it is still unclear which pathway ATRA induces the differentiation of solid cancers

Method used

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  • Pharmaceutical composition for removing residual liver cancer stem cells with combined use of arsenic trioxide and all-transretinoic acid and application thereof
  • Pharmaceutical composition for removing residual liver cancer stem cells with combined use of arsenic trioxide and all-transretinoic acid and application thereof
  • Pharmaceutical composition for removing residual liver cancer stem cells with combined use of arsenic trioxide and all-transretinoic acid and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Arsenic reduces the expression of PML protein in LCSCs:

[0024] 【Purpose】:

[0025] Need to specify As 2 o 3 Whether to down-regulate the expression of PML protein in LCSCs.

[0026] 【experimental method】:

[0027] (1) CD133 + CD13 + LCSCs sorting: HuH7 cells were cultured in DMEM with 10% FBS at 37°C, 5% CO 2 , incubator with saturated humidity, and when the confluence of the cells reached 70-80%, the cells were digested with 0.25% trypsin to make a single-cell suspension, and the cells were counted, washed once with PBS, put into 1.5ml EP tubes, and set blank Control tube, CD13, CD133, CD13-CD133 double antibody four groups, add 5μl antibody to each tube, incubate at 4°C in the dark for 30min, centrifuge at 700r / min for 5min, discard supernatant, wash 3 times with PBS, resuspend cells in 300μl PBS, Flow cytometry (BD FACSAria II and BD FACSCalibur) was used to detect the expression of CD13 and CD133 markers and sort LCSCs.

[0028] (2) Immunofluorescence anal...

Embodiment 2

[0038] Arsenic induces LCSCS differentiation:

[0039] (1) Arsenic (As 2 o 3 ) Down-regulate the expression of Oct4, Sox2, Klf4, the key genes of liver cancer stem cells

[0040] 【Purpose】:

[0041] As has been found 2 o 3 Higher than 0.8μg / ml inhibits the proliferation of liver cancer cells, and may induce apoptosis of liver cancer cells at high concentrations. Now we need to observe 0.5μg / ml As 2 o 3 CD133 + CD13 + Whether the functional characteristics of LCSCs and the expression of key genes have any effect, and whether LCSCs can be induced to differentiate.

[0042] 【experimental method】:

[0043] ① CD133 + CD13 + LCSCs sorting: Liver cancer tissues and HuH7 cells were cultured in 10% FBS in DMEM, 37°C, 5% CO 2 , incubator with saturated humidity, and when the confluence of the cells reached 70-80%, the cells were digested with 0.25% trypsin to make a single-cell suspension, and the cells were counted, washed once with PBS, put into 1.5ml EP tubes respectively...

Embodiment 3

[0104] ATRA downregulates PML protein expression in LCSCs:

[0105] 【Purpose】:

[0106] Since ATRA can induce the differentiation of various solid tumor cells including cancer stem cells and breast cancer cells, it is necessary to determine what molecular pathways actually induce cell differentiation or inhibit the proliferation of liver cancer cells, and whether they have common target molecules with arsenic PML protein.

[0107] 【experimental method】:

[0108] (1) CD133 + CD13 + LCSCs sorting:

[0109] Liver cancer cells were cultured in DMEM with 10% FBS at 37°C and 5% CO 2 , incubator with saturated humidity, and when the confluence of the cells reached 70-80%, the cells were digested with 0.25% trypsin to make a single-cell suspension, and the cells were counted, washed once with PBS, put into 1.5ml EP tubes, and set blank Control tube, CD13, CD133, CD13-CD133 double antibody four groups, add 5μl antibody to each tube, incubate at 4°C in the dark for 30min, centrifu...

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PUM

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Abstract

The invention discloses a pharmaceutical composition for removing residual liver cancer stem cells with combined use of arsenic trioxide As2O3 and all-transretinoic acid ATRA; an effective component is arsenic trioxide and all-transretinoic acid according to the final concentration of the total liquid amount. The invention also provides an application of the pharmaceutical composition in preparation of drugs for preventing and treating recurrence and metastasis of liver cancer after resection. The beneficial effects comprise that according to the pharmaceutical composition and the applicationthereof provided by the invention, arsenic trioxide is disclosed to have an effect of inducing differentiation of the liver cancer stem cells, and arsenic trioxide and all-transretinoic acid can inhibit PML proteins; moreover, arsenic trioxide and all-transretinoic acid can regulate down the expression of a key gene Oct4 of the PML downstream liver cancer stem cells respectively. A traditional wayof inducing apoptosis of liver cancer cells is broken through, and aiming at the residual liver cancer stem cells after resection of liver cancer, the differentiation of the liver cancer stem cells is induced by small-dose arsenic trioxide; with the combination of all-transretinoic acid and arsenic trioxide, the pharmaceutical composition can play a synergistic role, reduces the amount of arsenictrioxide and achieves the effect of eliminating the residual liver cancer stem cells.

Description

technical field [0001] The invention relates to the technical field of anticancer drugs, in particular to a pharmaceutical composition used in combination with arsenic and all-trans retinoic acid for eliminating residual liver cancer stem cells and its application. Background technique [0002] Hepatocellular carcinoma (Hepatocellar carcinoma, HCC) is the fifth most common malignant tumor in the world in terms of incidence, and the second in terms of mortality. my country is a high-incidence area of ​​HCC, accounting for almost 50% of the global incidence. HCC is closely related to hepatitis B virus infection and liver cirrhosis. In my country, hepatitis B virus infection accounts for 10% of the total population, and the incidence of HCC may remain high for a long time. When HCC is diagnosed, most of them are in the advanced stage, and the surgical resection rate is less than 50%. Even with surgical resection, it is easy to relapse and metastasis, and the 5-year survival ra...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K33/36A61P35/00A61P35/04A61K31/203
CPCA61K31/203A61K33/36A61P35/00A61P35/04A61K2300/00
Inventor 金世龙密雷王伟金榆凯
Owner 金世龙
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