SNP (Single Nucleotide Polymorphism) marker related to keratoconus genetic risk, application of SNP marker and keratoconus genetic risk assessment system
By constructing a multi-gene genetic risk scoring model containing 79 SNP sites, the problem of early keratoconus diagnosis was solved, efficient genetic risk assessment of the Chinese population was achieved, and personalized prevention and treatment were supported.
Patent Information
- Application Number
- CN202511171789.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-08-20
- Publication Date
- 2025-10-17
AI Technical Summary
Existing technologies make it difficult to effectively diagnose early-stage keratoconus, especially in the Chinese population due to the lack of a multi-gene genetic risk scoring model, which makes early diagnosis difficult and affects the prevention and treatment of the disease.
Using markers from 79 SNP loci, we construct a multi-gene genetic risk scoring model by detecting individual genotypes. Combined with Sanger sequencing, SNapshot genotyping, Taqman genotyping and other methods, we assess an individual's genetic risk of keratoconus and provide kits and assessment systems.
It has achieved high-accuracy and sensitive assessment of the genetic risk of keratoconus, which helps to identify high-risk groups at an early stage, conduct personalized diagnosis and treatment and intervention, and improve disease prevention effects.
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Figure CN120796465A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the field of biomedical technology, in particular to SNP markers related to the genetic risk of keratoconus and the application thereof and a keratoconus genetic risk assessment system. BACKGROUND
[0002] Keratoconus is a blinding eye disease characterized by corneal expansion, central or paracentral corneal stroma thinning, forward conical protrusion, and high myopia with irregular astigmatism. The prevalence of the disease is about 1.38 / 1000, and it usually occurs during adolescence. The disease progresses, and the visual acuity decreases severely in the late stage, which seriously affects the quality of life of patients and brings a heavy burden to families and society.
[0003] The main symptoms of keratoconus in the early stage are myopia and / or astigmatism in one or both eyes, and the myopia and astigmatism progress. In the middle and late stages, typical signs such as Fleischer ring, Vogt line, Munson sign, and central or inferior temporal thinning of the cornea can be observed under a slit lamp microscope. However, the diagnosis of early keratoconus is extremely difficult, especially in the very early stage. Current research shows that keratoconus has a family aggregation tendency, and genetic factors play an important role in the development of the disease. Multi-gene genetic risk score is a quantitative assessment of the genetic risk of complex diseases by considering multiple genetic variation sites in individuals, which has attracted widespread attention in the field of genetics. Currently, multi-gene genetic risk scores integrating multiple genetic variation information are widely used in diseases such as coronary heart disease and stroke. However, there is no report on the multi-gene genetic risk score of keratoconus in the Chinese population. Thin corneal thickness and steep corneal curvature are the main clinical features of keratoconus, so integrating multi-gene genetic risk scores related to central corneal thickness and corneal curvature is of great significance for the primary prevention of keratoconus. SUMMARY
[0004] To solve the above problems, the present application provides SNP markers related to the genetic risk of keratoconus and the application thereof and a keratoconus genetic risk assessment system.
[0005] To achieve the above-mentioned purpose, the present application provides the following technical solutions:
[0006] The present application provides a SNP marker related to the genetic risk of keratoconus, which comprises 79 SNP sites, and the information of the 79 SNP sites is shown in Table 1:
[0007] Table 1 Information of 79 SNP sites
[0008]
[0009]
[0010]
[0011] wherein the position is the physical position on the chromosome in the GRCh37 (hg19) reference genome to which the SNP site (the first base of the effect allele or the other allele) corresponds.
[0012] The present application provides a use of a reagent for detecting the SNP marker in the above technical solution in the preparation of a product for diagnosing keratoconus and / or evaluating genetic risk of keratoconus.
[0013] Preferably, the reagent is a reagent for detecting the genotype of the 79 SNP sites.
[0014] Preferably, the reagent comprises primers specific for amplifying the SNP marker or probes specific for binding the SNP marker.
[0015] Preferably, the method of detection comprises one or more of Sanger sequencing, SNapshot genotyping, Taqman genotyping and KASP genotyping.
[0016] Preferably, the product comprises a kit or an evaluation system.
[0017] The present application provides a system for evaluating genetic risk of keratoconus, comprising:
[0018] a sample collection module for collecting a sample to be tested of a person to be evaluated;
[0019] a genotyping module connected to the sample collection module for obtaining the genotype of the SNP site of the sample to be tested; the genotype of the SNP site is the genotype of the 79 SNP sites in the SNP marker described in the above technical solution;
[0020] a risk evaluation module connected to the genotyping module for evaluating the risk score of the sample to be tested; the evaluation comprises: obtaining the risk score of the sample to be tested by using formula I;
[0021]
[0022] wherein PRS j represents the risk score of the sample to be tested; j represents the sample to be tested; i represents the SNP site; G ij represents the number of effect alleles of the ith SNP carried by the sample to be tested, represented by 0, 1 and 2 in turn to represent no mutation, heterozygous mutation and homozygous mutation; Mj is the number of alleles contained in the PRS of the sample to be tested, i.e. the number of other alleles and effect alleles; S iis the effect value of the i th SNP, and specific values are shown in Table 2:
[0023] Table 2: Multi-gene genetic risk score site and effect value
[0024]
[0025]
[0026]
[0027] The result output module is connected with the risk assessment module, and is used for outputting the evaluation result of the to-be-evaluated person; the output is that the higher the risk score of the to-be-evaluated person is, the higher the risk of keratoconus is.
[0028] Preferably, the to-be-evaluated person includes Chinese.
[0029] Preferably, the to-be-evaluated sample includes peripheral blood.
[0030] Preferably, the genotyping module includes primers specific to the SNP markers or probes specific to the SNP markers.
[0031] Beneficial effects:
[0032] The SNP marker provided by the application is a SNP marker related to the genetic risk of keratoconus. The application finds SNP sites that are different (P<0.05) between the keratoconus group and the control group by screening and analyzing the SNP sites in 670 keratoconus patients and 4373 controls in the training set. Then, these sites are selected to construct a PRS model, and a keratoconus multi-gene genetic risk score model is constructed according to the effect value (lnOR) of different sites in the training set. Finally, 79 sites are retained, and a SNP marker related to the genetic risk of keratoconus is obtained. The SNP marker provided by the application can be used to evaluate the genetic risk of keratoconus, has high accuracy and sensitivity, and is helpful for the primary prevention of keratoconus. BRIEF DESCRIPTION OF DRAWINGS
[0033] In order to more clearly illustrate the technical solutions in the embodiments of the application or the prior art, the drawings needed in the embodiments will be briefly introduced as follows.
[0034] Figure 1 is the ROC curve in the training set;
[0035] Figure 2 is the ROC curve in the verification set;
[0036] Wherein, Sensitivity is sensitivity, sensitivity = true positive number / (true positive number + false negative number) x 100%; Specificity is specificity, specificity = true negative number / (true negative number + false positive number) x 100%. DETAILED DESCRIPTION
[0037] The present application provides a SNP marker related to the genetic risk of keratoconus, which includes 79 SNP sites, and the information of the 79 SNP sites is shown in Table 1.
[0038] The present application finds the SNP sites with differences (P<0.05) between the keratoconus group and the control group by screening and analyzing the SNP sites of 670 keratoconus patients in the training set and 4373 controls. Further, these sites are selected to construct a PRS model, and a keratoconus polygenic genetic risk score model is constructed according to the effect value (lnOR) of different sites in the training set. Finally, 79 sites are retained, and a SNP marker related to the genetic risk of keratoconus is obtained. By detecting these single nucleotide polymorphism sites, the genetic risk of keratoconus in the Chinese population can be evaluated, which has high accuracy and sensitivity, and is helpful for the primary prevention of keratoconus.
[0039] Based on the above advantages, the present application provides a reagent for detecting the SNP marker in the preparation of a product for diagnosing keratoconus and / or evaluating the genetic risk of keratoconus.
[0040] As an embodiment, the reagent is a reagent for detecting the genotype of the 79 SNP sites. As an embodiment, the reagent includes primers specific to the SNP marker or probes specific to the SNP marker. As an embodiment, the detection method includes one or more of Sanger sequencing, SNapshot genotyping, Taqman genotyping and KASP genotyping. As an embodiment, the product includes a kit or an evaluation system.
[0041] The present application provides a system for evaluating the genetic risk of keratoconus, comprising:
[0042] A sample collection module for collecting a sample to be tested of a person to be evaluated;
[0043] A genotyping module connected to the sample collection module for obtaining the SNP site genotype of the sample to be tested; the SNP site genotype is the genotype of the 79 SNP sites in the SNP marker described in the above technical solution.
[0044] The risk assessment module is connected with the genotyping module and is used for assessing the risk score of the sample to be tested; the assessment comprises: obtaining the risk score of the sample to be tested by using formula I.
[0045]
[0046] wherein PRS j represents the risk score of the sample to be tested; j represents the sample to be tested; i represents the SNP site; G ij represents the number of effect alleles of the ith SNP carried by the sample to be tested, and is represented by 0, 1 and 2 in sequence to represent no mutation, heterozygous mutation and homozygous mutation; Mj is the number of alleles contained in the PRS of the sample to be tested, i.e. the number of other alleles and effect alleles (if the individual to be tested detects the information of the 79 sites, then Mj is 79x2=158; if the individual to be tested only detects the information of 78 sites, then Mj is 78x2=156, and so on); S i represents the effect value of the ith SNP, and is specifically shown in Table 2;
[0047] The result output module is connected with the risk assessment module and is used for outputting the assessment result of the person to be assessed; the output is that the higher the risk score of the person to be assessed is, the higher the risk of keratoconus is.
[0048] As an implementation manner, the sample to be tested comprises peripheral blood. As an implementation manner, the person to be assessed is from a Chinese population.
[0049] As an implementation manner, the genotyping module comprises primers specifically amplifying the SNP markers or probes specifically binding to the SNP markers.
[0050] The system provided by the application has a curve under area of 0.906 in a training set and a curve under area of 0.853 in a verification set, which indicates that the system has a high genetic risk assessment capability for keratoconus.
[0051] In another aspect, the system provided by the application can be a computer storage medium storing computer program instructions, and the computer program instructions are executed to obtain the keratoconus risk assessment result of an individual based on the SNP site genotype of the individual to be tested.
[0052] In another aspect, the system provided by the application can be a computer device comprising a memory, a processor and a computer program stored in the memory and executable on the processor, and the processor executes the computer program to obtain the keratoconus risk assessment result of an individual based on the SNP site genotype of the individual to be tested.
[0053] The system provided by the present invention helps to identify people at high risk of keratoconus, and then provide personalized diagnosis, treatment and intervention for high-risk groups, providing more comprehensive, accurate and personalized technical support for risk assessment and prevention and control of keratoconus in my country.
[0054] To further illustrate the present invention, the SNP markers associated with the genetic risk of keratoconus, their applications, and the genetic risk assessment system for keratoconus provided by the present invention are described in detail below in conjunction with the examples and drawings, but they should not be construed as limiting the scope of protection of the present invention.
[0055] Example 1
[0056] 1. Study population
[0057] The present invention constructs and validates a polygenic genetic risk scoring model for keratoconus in the Chinese population in 958 patients with keratoconus and 6248 controls. The diagnosis of keratoconus is based on the following criteria: at least one of the following signs is positive in slit lamp examination (Fleischer ring, Vogt line, Munson sign, corneal scar); the central or paracentral cornea is dilated and thinned and protrudes forward in a cone shape; corneal topography examination reveals typical changes such as abnormal elevation of the anterior and posterior surfaces of the keratoconus. The present invention divides the 958 patients with keratoconus and 6248 controls into a training set and a validation set in a ratio of 7:3, of which 670 patients with keratoconus and 4373 controls are the training set, and 288 patients with keratoconus and 1875 controls are the validation set. The clinical characteristics of the training set study population are shown in Table 3, and the clinical characteristics of the validation set study population are shown in Table 4.
[0058] Table 3 Clinical characteristics of the keratoconus group and the control group in the training set
[0059] Parameters Keratoconus group (n=670) Control group (n=4373) Gender (male / female) 504 / 166 2133 / 2240 Age (years) 21.99±6.91 33.46±15.42 Central corneal thickness (pm) 436.80±59.40 - Mean corneal curvature (D) 53.22±8.52 -
[0060] Table 4 Clinical characteristics of the keratoconus group and the control group in the validation set
[0061] Parameters Keratoconus group (n=288) Control group (n=1875) Gender (male / female) 216 / 72 915 / 960 Age (years) 22.07±6.66 33.50±15.48 Central corneal thickness (pm) 435.73±55.73 - Mean corneal curvature (D) 53.46±8.22 -
[0062] 2. Selection of genetic variation sites
[0063] The whole genome association studies related to corneal thickness and corneal curvature were retrieved from the literature, and the genome-wide significant associations with corneal thickness and corneal curvature (P < 5 × 10 -8 ) genetic variation sites, and a total of 378 corneal thickness-related and 82 corneal curvature-related single nucleotide polymorphism sites were screened for the subsequent construction of multi-gene genetic risk scores.
[0064] 3. Genotyping
[0065] According to the principle of informed consent, the peripheral venous blood of the keratoconus patients and the control group individuals is collected, and then the genomic DNA of the research subjects is extracted. The qualified samples are subjected to DNA amplification, DNA fragmentation, chip hybridization, chip extension staining and chip scanning, and the SNP typing data of each sample is obtained. The genotyping results are obtained by the following steps: the treated chip is placed into a scanner, the fluorescence group of the single base extension product on the chip is excited by laser, the scanner obtains the fluorescence emitted by the fluorescence group, and a high-resolution picture is generated. The data obtained is directly imported into the Genome Studio software for analysis, so as to obtain the SNP typing data of each sample.
[0066] 4. Construction of keratoconus polygenic genetic risk score
[0067] The selected sites (378 corneal thickness related and 82 corneal curvature related single nucleotide polymorphism sites) are subjected to association analysis, and a total of 124 sites are different between the keratoconus group and the control group (P<0.05), and the results are shown in Table 5. Further, these sites are selected to construct the PRS model by PRSice software, the keratoconus polygenic genetic risk score model is constructed according to the effect value (lnOR) of different sites in the training set, and the optimal prediction model contains 79 sites (Table 2), and the model formula is:
[0068]
[0069] wherein j represents the individual to be tested, i represents the SNP site, S i is the effect value of the i th SNP, G ij represents the number of effect alleles carried by the individual to be tested, and 0, 1, 2 represent no mutation, heterozygous mutation and homozygous mutation in turn, and Mj is the number of alleles contained in the PRS of the individual to be tested. The effect value of each SNP is shown in Table 2. Then the ROC curve of the model is drawn, and the results are shown in Figure 1 , and the area under the curve is 0.906.
[0070] Table 5 Association analysis of SNP sites between keratoconus group and control group
[0071]
[0072]
[0073]
[0074]
[0075] Note: OR values in Table 1 are results of retaining three decimal places, and original data of OR values are used when calculating corresponding effect values (lnOR).
[0076] 5. Verification of the polygenic genetic risk score of keratoconus
[0077] The polygenic genetic risk score model constructed by the present application is verified in 288 keratoconus patients and 1875 controls, and the ROC curve of the polygenic genetic risk score model in the verification set is drawn, and the results are shown in Figure 2 , and the area under the curve is 0.853.
[0078] The area under the ROC curve greater than 0.9 indicates very high accuracy, and the area between 0.7 and 0.9 indicates high accuracy. The area under the curve of the polygenic genetic risk score model provided by the present application in the training set is 0.906, and the area under the curve in the verification set is 0.853, indicating that the model has high keratoconus discrimination ability.
[0079] Although the above embodiment describes the present application in detail, it is only a part of the embodiments of the present application, not all the embodiments, and other embodiments can be obtained according to the present embodiment without creativity, which all belong to the protection scope of the present application.
Claims
1. A SNP marker associated with the genetic risk of keratoconus, characterized in that: The SNP markers include 79 SNP sites, and the information of the 79 SNP sites is as follows: The position is the physical position of the SNP site on the corresponding chromosome in the GRCh37 reference genome.
2. Use of a reagent for detecting the SNP marker described in claim 1 in the preparation of a product for diagnosing keratoconus and / or assessing the genetic risk of keratoconus.
3. The use according to claim 2, characterized in that The reagent is a reagent for detecting the genotypes of the 79 SNP sites.
4. The use according to claim 2 or 3, characterized in that The reagents include primers that specifically amplify the SNP marker or probes that specifically bind to the SNP marker.
5. The use according to claim 2 or 3, characterized in that The detection method includes one or more of Sanger sequencing, SNapshot genotyping, Taqman genotyping and KASP genotyping.
6. The use according to claim 2 or 3, characterized in that The product includes a kit or an evaluation system.
7. A system for assessing genetic risk of keratoconus, characterized in that: include: A sample collection module is used to collect samples to be tested from the person to be evaluated; A genotyping module, connected to the sample collection module, for obtaining the SNP site genotype of the sample to be tested; the SNP site genotype is the 79 SNP site genotypes in the SNP marker of claim 1; A risk assessment module, connected to the genotyping module, is used to assess the risk score of the sample to be tested; the assessment includes: obtaining the risk score of the sample to be tested using Formula I; Among them, PRS j represents the risk score of the sample to be tested; j represents the sample to be tested; i represents the SNP site; G ij Refers to the number of effect alleles of the i-th SNP carried by the sample to be tested, with 0, 1, and 2 representing no mutation, heterozygous mutation, and homozygous mutation respectively; Mj is the number of alleles contained in the PRS of the sample to be tested, that is, the number of other alleles and effect alleles; S i It refers to the effect value of the i-th SNP, as follows: The result output module is connected to the risk assessment module and is used to output the assessment results of the person to be assessed; the output is: the higher the risk score of the person to be assessed, the higher the risk of keratoconus.
8. The system according to claim 7, characterized in that The sample to be tested includes peripheral blood.
9. The system according to claim 7, wherein: The people to be assessed include Chinese.
10. The system according to claim 7, wherein: The genotyping module includes: a primer for specifically amplifying the SNP marker or a probe for specifically binding to the SNP marker.