Combined marker for early diagnosis of liver cancer and its application
A marker and liver cancer technology, applied in the field of biomedicine, can solve the problems of lack of effective therapeutic targets, difficulties in early diagnosis, high recurrence and metastasis rate, and achieve the effect of avoiding tissue biopsy, improving early screening sensitivity, and improving sensitivity
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Embodiment 1
[0070] Example 1. Establishment of a method for detecting liver cancer
[0071] 1. Markers for liver cancer detection
[0072] The inventors of the present invention combined various databases and comprehensive clinical information, designed probes to enrich the methylation sites of related genes through large-scale screening, and then compared the cfDNA 3-6 months before the diagnosis of liver cancer through library construction and sequencing. Differences in methylation levels between samples and cfDNA samples from non-HCC patients to obtain markers for liver cancer detection. The markers include methylation levels of any 6 or more of the following 17 genes and / or mutations in at least one of the two genes TERT and CTNNB1:
[0073] AK055957, DAB2IP, GRASP, PPFIA1, PSD4, TBX15, MKL1, DCDC2, BCL2L11, SEMA4D, PROX1, FAM78A, PTPN7, AXIN1, SHH, SR, and CYP1B1.
[0074] The positions of these genes (reference genome GRCh38 / hg38) are shown in Table 1, wherein, 1) TBX15, MKL1, DCD...
Embodiment 2
[0125] Example 2. Using the 7 markers of the present invention to detect samples
[0126] The selected samples were 12 clinical liver cancer tissue samples and 12 healthy human leukocyte samples.
[0127] The inclusion criteria of 12 patients with clinical liver cancer were as follows: the diagnosis of liver cancer was confirmed by pathological examination of histological sections.
[0128] The inclusion criteria of 12 healthy people were: healthy non-HCC patients.
[0129] The seven markers are: MKL1, DCDC2, BCL2L11, CYP1B1, PTPN7, AXIN1 and SEMA4D gene methylation.
[0130] 1. The test sample is first passed through Qiagen's Genomic DNA was extracted using the Mini Kit kit according to the instructions in the instructions. The OD260 / 280 of DNA should be between 1.8-2.1 as determined by Nanodrop from ThermoFisher.
[0131] 2. Methylation detection was carried out to the extracted DNA, and according to the method of step 2 of Example 1, transformation, pre-amplification a...
Embodiment 3
[0137] Example 3. Using the 10 markers of the present invention to detect samples
[0138] The selected samples are the same as those in Example 2.
[0139] 10 markers: FAM78A, MKL1, DCDC2, BCL2L11, PTPN7, AXIN1, PROX1, AK055957, PPFIA1 and PSD4 gene methylation.
[0140] The detection steps are the same as those in Example 2. According to the methylation detection method in Step 2 of Example 1, the methylation of ten genes of FAM78A, MKL1, DCDC2, BCL2L11, PTPN7, AXIN1, PROX1, AK055957, PPFIA1 and PSD4 were detected. , and calculate the normalized value L of these ten genes respectively 目的基因 , the normalized value L for these ten genes 目的基因 Sum, get HCCscan, HCCscan=L FAM78A +L MKL1 +L DCDC2 +L BCL2L11 +L PTPN7 +L AXIN1 +L PROX1 +L AK055957 +L PPFIA1 +L PSD4 , the results of each sample are shown in Table 11.
[0141] Table 11. Test Results
[0142] sample name sample type HCCscan sample name sample type HCCscan T25 cancer tissue 105.4...
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