Serum miRNA marker for OPLL diagnosis and application thereof
Inventors
Liu, Yang • Xu, Chen • Zhang, Hao • Yuan, Wen • Cao, Peng • WU, Huiqiao • Chen, Yuanyuan • SHEN, Xiaolong
Assignees
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Publication Number
US-11124832-B2
Publication Date
2021-09-21
Expiration Date
Abstract
A serum microRNA (miRNA) marker suitable for early screening and diagnosis of ossification of posterior longitudinal ligament (OPLL) and its application in the diagnostic reagent or kit for the OPLL. Biomarker miRNA-563, miRNA-196b, miRNA-10a and miRNA-129 have high diagnostic value for OPLL, and the development and application of the related serum miRNA biomarker detection reagent kit. It can be applied in the screening of ossification of posterior longitudinal ligament disease, supporting the diagnosis of OPLL more quickly and accurately, evaluate the patient's ossification condition, and may lay down the foundation for improving clinical therapeutic effect.
Core Innovation
The patent discloses serum microRNA (miRNA) biomarkers for early screening and diagnosis of ossification of the posterior longitudinal ligament (OPLL). The biomarkers include miRNA-563, miRNA-196b, miRNA-10a, and miRNA-129, which are positively correlated with OPLL risk as described in the provided summary content.
The disclosed approach involves measuring expression levels of the specified miRNAs in a biological sample and using PCR primers corresponding to SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. A diagnostic risk value is calculated from miRNA copy numbers, and the risk value is used as an indicator of high risk when the risk value is greater than 0.
The patent further describes diagnostic reagents and kits for detecting the miRNAs, preferably using Real-time RT-PCR/qPCR. Experimental verification is reported using sequencing and qPCR across multiple sample sets, including elevated serum levels in OPLL versus controls and improved diagnostic performance when using a combined analysis for the four miRNAs.
Claims Coverage
The provided independent claim covers a diagnosing method for OPLL based on measuring expression levels of four specified miRNAs using specific PCR primers, followed by calculating a composite risk value from miRNA copy numbers. The risk value threshold defines a high-risk result, and the dependent claims mainly refine the biological sample scope and/or narrow the measuring step to Real-time PCR.
Measuring miRNA expression with specified PCR primers
A method of diagnosing ossification of the posterior longitudinal ligament by measuring expression levels of at least one of miRNA-563 (SEQ ID NO: 1), miRNA-196b (SEQ ID NO: 2), miRNA-10a (SEQ ID NO: 3) and miRNA-129 (SEQ ID NO: 4) in a biological sample by using at least one of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8 as PCR primers, respectively.
Composite miRNA copy-number risk value for OPLL diagnosis
Calculating a risk value of the composite expression based on miRNA-563, miRNA-196b, miRNA-10a, and miRNA-129 copy numbers per specified serum volumes, where the risk value is greater than 0 indicating a high risk of ossification of the posterior longitudinal ligament.
Across the provided claims, the core inventive coverage is directed to measuring specified OPLL-associated miRNAs using the specified PCR primers and calculating a composite risk value from miRNA copy numbers with the threshold that risk value greater than 0 indicates high risk.
Stated Advantages
Improved diagnostic performance when using the combined analysis of the four miRNAs compared with individual miRNAs, including higher specificity as described in the provided summary content.
Documented Applications
Diagnostic reagents and kits for diagnosing ossification of the posterior longitudinal ligament using detection of serum miRNA biomarkers, preferably using Real-time RT-PCR/qPCR as described in the provided summary content.
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