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Synergetic Application of nicotinamide phosphoribosyltransferase (NAMPT) depressor and NQO1 substrate to treatment of non-small cell lung cancer

A technology of non-small cell lung cancer and inhibitors, applied in drug combinations, medical preparations containing active ingredients, pharmaceutical formulas, etc.

Inactive Publication Date: 2014-08-06
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There is currently no effective and low-toxicity treatment

Method used

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  • Synergetic Application of nicotinamide phosphoribosyltransferase (NAMPT) depressor and NQO1 substrate to treatment of non-small cell lung cancer
  • Synergetic Application of nicotinamide phosphoribosyltransferase (NAMPT) depressor and NQO1 substrate to treatment of non-small cell lung cancer
  • Synergetic Application of nicotinamide phosphoribosyltransferase (NAMPT) depressor and NQO1 substrate to treatment of non-small cell lung cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1.PCR experiment

[0021] Experimental Materials:

[0022] Human non-small cell lung cancer A549 cells were purchased from ATCC at 37°C, 5% CO 2 Under normal conditions, culture medium was RPMI-1640 (Gibco) containing 10% calf serum (PAA); fetal bovine serum (Fetal bovine serum) was purchased from Hyclone (Logan, Utah, USA); trypsin (Trypsin) was purchased from in Amersco (Solon, Ohio, USA); penicillin, streptomycin, dimethylthiodiphenyltettrazole (MTT), and GAPDH antibodies were purchased from Nanjing Shengxing Bioengineering Company (Jiangsu); Trizol total RNA extraction reagent , RT-PCR kit, and Real-time PCR Master Mix (SYBR Green) were purchased from Takara (Dalian Baobiology).

[0023] experimental method:

[0024] A549 cells were seeded in 6-well plates, and the treatment time was 0h, 2h, 4h, 8h, 12h, 24h; per 1×10 6 Add 1mL Trizol to the cells, blow the liquid with a 1mL pipette until the liquid is clear and free of cell clumps, and transfer it i...

Embodiment 2

[0029] Embodiment 2.MTT experiment

[0030] A549 cells were seeded in a 96-well plate, cultured and grown to 80% full, given FK866 10 nM, and treated with different concentrations of Tanshinone IIA (0, 0.4, 1, 2, 4, 10, 20, 40 μM) for 72 hours after 1 hour; or β-La Paquinone (0, 1.25, 2.5, 5, 10, 20 μM) was treated for 2 hours, and then the drug-free medium was replaced for 24 hours. Replace the serum-free medium and add 20 μL (5 mg / mL) MTT to each well, incubate in a carbon dioxide incubator for 4 hours, remove the MTT solution, add 150 μL of DMSO to dissolve the crystals (37°C shaker, 50r / min), take it out after 10 minutes and place in The absorbance was measured in a microplate reader (measurement wavelength 570nm, reference wavelength 630nm). The cell survival rate of the administration group was calculated by the following formula:

[0031] Survival rate = absorbance of treatment group / absorbance of control group * 100%

Embodiment 3

[0032] Example 3.LC-MS n Detection of intracellular NAD + Level

[0033] A549 cells were inoculated into 6-well plates, cultured and grown to 80% confluence, treated with FK866 10nM, 1 hour later with 40 μM Tanshinone IIA for 48 hours; or 20 μM β-lapachone for 24 hours. Discard the medium, wash the cells with normal saline, add 1ml of 80% methanol containing 50ng / ml 2-chloroadenosine (internal standard) to each well, place at -80°C for 20 minutes, scrape the cells in an ice bath and transfer them to EP tubes, and sonicate Break up, centrifuge at 14000rpm for 5 minutes, transfer the supernatant to a new EP tube, evaporate to dryness, reconstitute and inject in 100μl ultrapure water.

[0034]Chromatographic conditions: the chromatographic column is an amide column (3x100mm i.d., 5 μm, Chrom-Matrix Inc), and the column temperature is 40°C. Mobile phase: the aqueous phase (A) is an aqueous solution containing 5mM ammonium acetate, the organic phase (B) is methanol, the column t...

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PUM

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Abstract

The invention relates to a combined drug, comprising a certain amount of nicotinamide phosphoribosyltransferase (NAMPT) depressor and a certain amount of NQO1 substrate. The NAMPT depressor and the NQO1 substrate have synergistic effects in the anti-tumor process, and the action of the composition in the case of combined utilization of the NAMPT depressor and the NQO1 substrate is superior to the superposition when all the drugs are independently used. The combined drug can be applied to treatment of the non-small cell lung cancer and other cancers. Therefore, a novel treatment method for treating the non-small cell lung cancer is provided by the invention.

Description

technical field [0001] The invention relates to the fields of natural medicine and pharmacy, in particular to the use of a NAMPT inhibitor combined with an NQO1 substrate to prepare a drug for treating tumors. More specifically, it relates to the use of NAMPT inhibitors and NQO1 substrates in synergistic treatment of non-small cell lung cancer. Background technique [0002] Non-small cell lung cancer is the malignant tumor with the highest morbidity and mortality, ranking first among the causes of death caused by tumors. There is currently no effective and low-toxicity treatment. [0003] Nicotinamide adenine dinucleotide nicotinamide adenine dinucleotide abbreviation NAD+. It can accept a hydrogen atom and an electron from the substrate in the oxidation-reduction process and become a reduced form, playing a central role in the electron transfer process of the respiratory chain. In recent years, the role of NAD+ in maintaining cellular metabolic activity, cellular stress ...

Claims

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Application Information

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IPC IPC(8): A61K45/06A61K31/58A61K31/352A61K31/4545A61P35/00
Inventor 王广基郝海平刘慧颖李清然程学芳
Owner CHINA PHARM UNIV
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