Tumor radiosensitizers containing eEF2K inhibitors
A radiosensitizer and inhibitor technology, applied in the clinical field, can solve the problems of target-specific toxicity, low radiation dose, and increase the curative effect of cancer radiotherapy, and achieve the goals of reducing complications, low radiation dose, and reducing the impact Effect
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Embodiment 1W
[0024] Example 1 Western Blot detection of the effect of NH-125 on eEF2K targeting inhibition of breast cancer cells
[0025] Two breast cancer cells MCF-7 and MDA-MB-231 were pretreated with different concentrations of NH-125 for 24 hours, then lysed with RIPA containing 10% PMSF, collected after shaking on ice for 15 minutes, centrifuged at 12000 rpm for 15 minutes, protein BCA After the kit is quantified, add 5X loading buffer and cook for 5 minutes to denature. Use 10% SDS-PAGE gel for 80V electrophoresis for one hour, then turn to 120V for another hour, turn PVDF membrane at 250mA for 100 minutes, block with 5% milk for 1 hour, incubate with primary antibody at 4°C overnight, wash 3 times with TBST, 15 minutes each time , incubated with secondary antibody for 1 hour, washed three times with TBST, each time for 15 minutes, and developed exposure.
[0026] The result is as figure 1 shown, from figure 1 It can be seen that the expression of eEF2K was significantly inhibit...
Embodiment 2C
[0027] Example 2 The CCK8 assay detects the effect of NH-125 on the proliferation of breast cancer cells:
[0028] Breast cancer cells MCF-7 and MDA-MB-231 were evenly spread in a 96-well plate at a density of 5000 cells / well overnight, and treated with different concentrations of NH-125 for 24 hours and 48 hours the next day, and then 10% The CCK8 was incubated at 37°C for 1.5h, and then the OD value was measured with a spectrophotometer at a wavelength of 450nm to calculate the cell proliferation.
[0029] The result is as figure 2 shown, from figure 2 It can be seen that with the increase of NH-125 dose, the cell viability decreased significantly.
Embodiment 3
[0030] Example 3 Cloning formation experiment to detect the influence of NH-125 on radiosensitivity of breast cancer cells:
[0031] Breast cancer cells MCF-7 and MDA-MB-231 were planted in six-well plates at different densities, and treated at concentrations of 0.1 μM and 0.25 μM for 24 h after cell attachment, and DMSO was used as a control. Then the cells were irradiated with 6MV X-rays at a dose rate of 4.5Gy per minute at 0, 2, 4, 6, and 8Gy, respectively, and then the cells were cultured for two weeks, fixed with methanol for 30 minutes, washed with running water three times, and stained with Giemsa stain. After 30 minutes, count the number of clones with more than 50 cells.
[0032] Table 1 Effect of NH-125 on radiosensitivity of breast cancer cells MCF-7
[0033]
[0034] Table 2 Effect of NH-125 on radiosensitivity of breast cancer cells MDA-MB-231
[0035]
[0036] The result is as image 3 , Table 1 and Table 2, from image 3 , Table 1 and Table 2, it can ...
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