Method for determining risk of urothelial carcinoma

A urothelial and determination method technology, which is applied in biochemical equipment and methods, microbial measurement/testing, DNA/RNA fragments, etc., can solve problems such as complicated measurement methods, difficult medical implementation, and unclear CpG sites. Achieve the effect of improving QOL and survival rate, high sensitivity and specificity

Inactive Publication Date: 2020-10-30
KEIO UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, in this method, the average amount of the DNA methylation rate of multiple CpG sites on the BAC clone is measured by hybridization of the entire BAC, and the CpG site with high diagnostic accuracy is unclear. In addition, the measurement method is complicated and difficult. medical implementation

Method used

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  • Method for determining risk of urothelial carcinoma
  • Method for determining risk of urothelial carcinoma
  • Method for determining risk of urothelial carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0151] [Example 1 Detection of methylation of CpG sites in cancerous tissues]

[0152] (Infinium (registered trademark) measurement)

[0153] The DNA methylation status of 485,764 CpG sites was analyzed at a resolution of one CpG site using the Infinium (registered trademark) HumanMethylation 450k Bead Chip (manufactured by Illumina). It is generally considered that the Infinium (registered trademark) HumanMethylation450k Bead Chip (manufactured by Illumina) covers 99% of RefSeq genes and 96% of CpG islands. More specifically, for the purpose of comprehensively analyzing DNA methylation status, analysis targets include promoter region, 5' untranslated region, first exon, intragene, 3' untranslated region, It is generally considered that 99% of the reference sequence genes are included.

[0154] The bisulfite-treated DNA was subjected to genome-wide amplification using the Infinium (registered trademark) assay kit (manufactured by Illumina) (see Bibikova, M. et al., Epigenomi...

Embodiment 2

[0164] [Example 2 Evaluation of markers for urothelial carcinoma]

[0165] Five genes (TENM3, HOXC4, TLR1, CPVL, and PRDM16) containing CpG sites with an AUC of 0.95 or more in the ROC curve prepared in Example 1 were selected, and the CpGs of these genes were confirmed by MassARRAY (registered trademark) or pyrosequencing DNA methylation levels in loci.

[0166] (MassARRAY(registered trademark) Act)

[0167] The MassARRAY (registered trademark) method is to amplify the bisulfite-treated DNA and transcribe it into RNA, and then use RNAase to perform base-specific cleavage, and then use a mass spectrometer to detect methylated DNA fragments and unmethylated DNA fragments. A method for the difference in molecular weight of DNA fragments.

[0168] Primers for MassARRAY were designed for the CpG site of the TENM3 gene using EpiDesigner (manufactured by SEQUENOM, primer design software for MassARRAY). Combinations of 3 types of DNA polymerases and 4 levels of annealing temperatu...

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Abstract

Provided is a method for determining the risk of urothelial carcinoma on the basis of DNA methylation level. A method for determining the risk of canceration of a urothelial tissue, said method comprising: detecting the CpG site DNA methylation level of at least one gene selected from the group consisting of TENM3, HOXC4, TLR1, CPVL and PRDM16 in genomic DNA derived from a urothelial cell or a tissue containing the same; and determining the risk of canceration of the urothelial tissue depending on the DNA methylation level thus detected.

Description

technical field [0001] The present invention relates to a method for determining the risk of urothelial carcinoma. Background technique [0002] Urothelial carcinoma is characterized clinically and pathologically by multiple temporal and spatial lesions, that is, the occurrence of lesions in multiple locations or in a wide range of simultaneous and metachronous lesions in the renal pelvis, ureter, and bladder. As a striking example of spatial multiples, there is high atypia of squamous intraepithelial carcinoma that spreads widely throughout the urinary tract from the renal pelvis to the urethra. Highly atypia squamous intraepithelial carcinoma is considered to be the precursor lesion of nonpapillary (nodular) invasive urothelial carcinoma with high malignancy and poor prognosis. In contrast, superficial papillary urothelial carcinoma with a low degree of atypia has a good prognosis despite frequent recurrences (metachronous multiple). However, the repeated recurrence of p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/154
Inventor 金井弥荣新井惠吏
Owner KEIO UNIV
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