Rs62226057-related SNP marker combination for evaluating high myopia risk based on ocular axis and application of rs62226057-related SNP marker combination

Abstract

The invention discloses an rs62226057 related SNP marker combination for evaluating a high myopia risk based on an ocular axis and application of the rs62226057 related SNP marker combination, and relates to the field of biological medicines. According to the invention, through large-scale queue analysis, a group of SNP marker combination based on the length of the ocular axis is screened and verified, and the key site in the SNP marker combination is rs62226057. The invention further provides a method, a system and equipment for evaluating the high myopia risk based on the SNP marker combination, an important tool is provided for identification and accurate intervention of people with the high myopia risk, and the method, the system and the equipment have wide clinical application value.

Description

Technical Field

[0001] This invention relates to the field of biomedicine, specifically to a combination of rs62226057-related SNP biomarkers for assessing the risk of high myopia based on axial length and their application. Background Technology

[0002] High myopia (HM), a key factor leading to irreversible visual impairment, presents a complex and not yet fully understood genetic basis in the Chinese population. Currently, high myopia is usually defined as an equivalent spherical power equal to or less than -6.00 diopters (D), or an axial length greater than 26 mm. This condition is closely associated with a series of serious vision-threatening fundus complications, such as myopic choroidal neovascularization (mCNV), retinal detachment (RD), and myopic macular degeneration (MMD). These complications can severely impair visual function, potentially leading to irreversible blindness and significantly impacting patients' quality of life. Notably, the prevalence of HM among adolescents aged 16-18 is showing a significant upward trend, projected to rise from 7.3% in 2001 to 22.1% in 2050, highlighting the severity of the HM prevention and control situation.

[0003] However, current research still faces significant challenges in identifying reliable and universally applicable risk biomarkers for high myopia. Existing biomarkers are limited in number and effect size, and exhibit poor reproducibility across different populations, resulting in a lack of effective biomarker systems for early warning or individualized assessment in clinical practice. This scarcity of high myopia risk biomarkers not only limits the accurate identification of high-risk individuals but also hinders in-depth analysis of disease mechanisms and the development of targeted intervention strategies. Given the significant genetic heterogeneity and phenotypic diversity of high myopia, relying solely on traditional diagnostic classifications is insufficient to capture its complex biological basis. Summary of the Invention

[0004] To address the shortcomings of existing technologies, this invention provides a combination of rs62226057-related SNP biomarkers for assessing the risk of high myopia based on axial length and their applications.

[0005] To achieve the above-mentioned objectives, the present invention provides the following technical solution: The first aspect of the present invention provides a method for constructing a model to assess the risk of high myopia based on axial length, the method comprising: obtaining axial length data of highly myopic individuals and non-highly myopic individuals, and constructing a high myopia risk model based on the genotype of SNP markers.

[0006] Furthermore, the high myopia risk model is constructed using algorithms, including one or more of the following: random forest model, Cox regression model, principal component analysis, deep neural network, generalized linear model, logistic regression analysis, LASSO regression analysis, nearest neighbor analysis, support vector machine, and neural network model.

[0007] In some embodiments, the method for constructing the high myopia risk model is known to those skilled in the art, and the steps of associating SNP locus genotypes with a certain probability or risk can be implemented and realized in different ways. Preferably, the results of SNP locus genotype determinations are mathematically combined, and the scores of different genotypes are associated with a fundamental risk assessment problem. The results of SNP locus genotype determinations can be combined using any suitable existing technical mathematical method, and the high myopia risk model can be constructed using algorithms.

[0008] Furthermore, the SNP markers are rs62226057: T>C, rs11582005: G>A, rs8191982: G>A, rs199649451: C>T, rs2257107: T>A, rs142759452: C>A, rs6423208: A>G, rs80046353: C>G, rs10932037: C>T, rs3792268: C>A, rs7631705: T>C, rs678690: C>T, rs7507: G>A, rs117234103: G>C, rs31315: T>C, rs77349207: T>C, rs611580: G>C, rs520589: C>T, rs164544: C>T, rs11551053: T>C, rs663227: T>C, rs3823465: A>G, rs2274954: T>C, rs142475387: A>G, rs12678121: C>T, rs80120341: C>T, rs1049832: A>G, rs67210953: G>A, rs4579578: G>T, rs1255411: C>T, rs200502145: G>A, rs56139616: G>A, rs34269909: T>A, rs35607186: T>C, rs34107711: A>G, rs61738000: C>T, rs1356428: A>T, rs76634080: C>G, rs79983883: C>A, rs7138453: T>C, rs59836757: C>T, rs3816503: G>T, rs7306824: G>A, rs79941046: A>G, rs114131205: C>T, rs2075260: G>A, rs2305901: G>A, rs78569455: G>C, rs2876702: T>C, rs3742137: C>T, rs9577407: G>A, rs8017100: T>C, rs2289818: G>C, rs55721315: G>T, rs143952603: G>A, rs8052283: G>A, rsrs6513497: T > G; rs817330: C > T; rs12483205: A > G combination.

[0009] A second aspect of the present invention provides a kit for assessing the risk of high myopia based on axial length, the kit comprising reagents for detecting SNP locus genotypes by one or more methods selected from nucleic acid hybridization technology, nucleic acid amplification technology, and sequencing technology, wherein the SNP locus is the SNP marker described in the first aspect.

[0010] In some embodiments, the reagent is a probe that specifically recognizes the genotype of the SNP site; or, the reagent is a primer that specifically amplifies the gene fragment containing the SNP site.

[0011] In this invention, the term "probe" refers to a molecule capable of selectively binding to a specifically intended SNP site, such as a nucleotide transcript or protein encoded by an intrinsic gene or corresponding to an intrinsic gene. Probes can be synthesized by those skilled in the art or can be derived from suitable biopreparations. Probes can be specifically designed to be labeled. Examples of molecules that can be used as probes include, but are not limited to, RNA, DNA, proteins, antibodies, and organic molecules.

[0012] In this invention, the term "primer" refers to a single-stranded polynucleotide capable of hybridizing with nucleic acids and allowing the polymerization of complementary nucleic acids (generally by providing a free 3'-OH group).

[0013] In some embodiments, the kit may further include one or more of the following: reagents for processing samples, standards, calibrators, buffer solutions, and instructions.

[0014] A third aspect of the present invention provides the application of a reagent for detecting the SNP markers described in the first aspect in the preparation of a product for assessing the risk of high myopia, said product including a reagent kit, a chip, a membrane strip, a system, an apparatus, and a storage medium.

[0015] In the context of this invention, the term "sample" as used refers to a composition obtained from or derived from a subject (e.g., an individual of interest) that contains cells and / or other molecular entities to be characterized and / or identified based on, for example, physical, biochemical, chemical, and / or physiological characteristics. For instance, a sample refers to any sample derived from a subject of interest that is expected or known to contain cells and / or molecular entities to be characterized.

[0016] In some embodiments, the sample comprises nucleic acids from the subject.

[0017] Furthermore, the sample was peripheral blood.

[0018] The term "chip," also known as an "array," refers to a solid support containing linked nucleic acid or peptide probes. An array typically contains a variety of different nucleic acid or peptide probes attached to a substrate surface at different known locations. Arrays can contain flat surfaces or can be nucleic acids or peptides on beads, gels, polymer surfaces, fibers such as optical fibers, glass, or any other suitable substrate. Arrays can be packaged in ways that allow for diagnostic or other manipulation of a fully functional device.

[0019] A "microarray" is a hybridization array element arranged in an ordered manner on a matrix, such as a polynucleotide probe (e.g., an oligonucleotide) or a binder (e.g., an antibody). The matrix can be a solid matrix, such as a glass or silica slide, beads, fiber optic adhesive, or a semi-solid matrix, such as a nitrocellulose membrane. The nucleotide sequence can be DNA, RNA, or any arrangement thereof.

[0020] Furthermore, the chip includes at least one of microfluidic chip, microarray chip, fiber optic microbead chip, liquid phase chip, and in-situ synthesis chip.

[0021] In this invention, the term "microfluidic chip" refers to a chip that can manipulate microfluidics on a chip to carry out various functions of conventional physical, chemical or biological experiments. The channel size on the chip can be on the order of micrometers (μm) or even nanometers (nm).

[0022] In this invention, the "membrane strip" is also called a "nucleic acid membrane strip", which includes a substrate and an oligonucleotide probe immobilized on the substrate; the substrate can be any substrate suitable for immobilizing oligonucleotide probes, including nylon membrane, nitrocellulose membrane, polypropylene membrane, glass slide, silicone wafer, micro magnetic beads, but is not limited thereto.

[0023] In this invention, the term "kit" includes reagents for detecting the SNP markers described in the first aspect of the invention, and one or more substances selected from the group consisting of: containers, instructions for use, positive controls, negative controls, buffers, auxiliaries, or solvents. Components of the kit may be packaged in an aqueous medium or in lyophilized form. Suitable containers in the kit typically include at least one vial, test tube, long-necked flask, PET bottle, syringe, or other container in which one component can be placed, and preferably appropriately aliquoted. When more than one component is present in the kit, the kit will also typically include a second, third, or other additional container in which the additional components are placed separately. However, different combinations of components may be contained in a single vial. The kit of the present invention will also typically include a container for containing the reactants, sealed for commercial sale. Such a container may include injection-molded or blow-molded plastic containers in which the desired vials can be retained.

[0024] A fourth aspect of the present invention provides a system for assessing the risk of high myopia based on axial length, the system comprising the following modules: The acquisition module is configured to acquire SNP marker data described in the first aspect from the sample; The processing module is configured to input the SNP marker data described in the first aspect of the sample into the high myopia risk model constructed by the method described in the first aspect for analysis, and obtain the analysis results; The output module is configured to output analysis results.

[0025] This invention provides a system programmed to implement the methods described herein. The system is programmed or otherwise configured to analyze sequence data and construct gene expression matrices. The system can control various aspects of sequence analysis according to the invention, such as, for example, matching data against known sequences. The system can be a user's electronic device or a computer system remotely located relative to that electronic device. The electronic device can be a mobile electronic device.

[0026] A fifth aspect of the present invention provides a computer device for assessing the risk of high myopia, the computer device including a memory and a processor, the memory being used to store computer programs.

[0027] The processor executes a computer program, which, when executed, implements the following method: Data is acquired to obtain SNP marker data as described in the first aspect of the sample; The data is processed to input the SNP marker data described in the first aspect of the sample into the high myopia risk model constructed by the method described in the first aspect for analysis, and to obtain the analysis results. Output results, used to output analysis results.

[0028] It should be understood that the systems, devices, and methods described in this invention can be implemented in other ways. For example, the system embodiments described above are merely illustrative. For instance, the division of modules is only a logical functional division, and in actual implementation, there may be other division methods. For example, multiple modules or components may be combined or integrated into another system, or some features may be ignored or not executed. Furthermore, the coupling or direct coupling or communication connection shown or discussed may be indirect coupling or communication connection through some interfaces, devices, or modules, and may be electrical, mechanical, or other forms.

[0029] A sixth aspect of the present invention provides a computer program product, including a computer program for assessing the risk of high myopia, wherein the computer program, when executed by a processor, implements the following method: Data is acquired to obtain SNP marker data as described in the first aspect of the sample; The data is processed to input the SNP marker data described in the first aspect of the sample into the high myopia risk model constructed by the method described in the first aspect for analysis, and to obtain the analysis results. Output results, used to output analysis results.

[0030] Advantages and benefits of this invention: This invention, through large-scale cohort analysis, screened and validated a combination of SNP markers based on axial length, with the key locus being rs62226057. The SNP marker combination based on this previously undisclosed locus can accurately assess the risk and susceptibility to high myopia, and has broad application value. Attached Figure Description

[0031] Figure 1 Manhattan plot results for the key site WNT7Brs62226057 in a high myopia dataset.

[0032] Figure 2 The ROC curves were calculated for different thresholds of the filter set, where A is 27 mm, B is 28 mm, and C is 29 mm.

[0033] Figure 3 ROC curves calculated for different thresholds on the validation set, where A is 27 mm, B is 28 mm, and C is 29 mm.

[0034] Figure 4 The ROC curves are calculated for different thresholds of the sampled set, where A is 27 mm, B is 28 mm, and C is 29 mm.

[0035] Figure 5 The graph shows the ROC curves calculated for different thresholds on the validation set after sampling, where A is 27 mm, B is 28 mm, and C is 29 mm. Detailed Implementation

[0036] To enable those skilled in the art to better understand the present invention, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments. Obviously, the described embodiments are merely some embodiments of the present invention, and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0037] Example I. Experimental Methods 1. Study Design and Participants: Participants in this study were from the Myopia-Related Genetics and Intervention Consortium (MAGIC) project. Some patients underwent ophthalmological examinations, including refractive error measurements and ocular biometrics. Subsequent analyses used corneal curvature (K) and axial length (AL) as the primary endpoints. High myopia was defined based on the worst-case axial length (WAL > 26.0 mm). This study was approved by the Eye Hospital of Wenzhou Medical University and conducted in accordance with the principles of the Declaration of Helsinki. All participants provided written informed consent.

[0038] 2. Exclusion Criteria for Participants: In the initial stage of data processing, individuals lacking valid identification or an AL (Audiovisual Length) were excluded. Participants who had previously undergone ophthalmic surgery were also excluded. Furthermore, participants were excluded if the axial length of the worst eye was less than or equal to 26.0 mm.

[0039] 3. Genotypic Data and Quality Control: All genotypic data were retrieved from the patient database collected by the MAGIC project and filtered for variants using a standardized approach. Then, the corresponding genotypes for the five ocular phenotypes were extracted. Second-stage quality control for variant levels included the following exclusion steps: recall <90%, Hardy-Weinberg equilibrium p <1×10⁻⁶. 6 The minor allele frequency (MAF) is <0.01.

[0040] 4. Biomarker Screening and Validation: The collected patient samples were randomly divided into two high myopia datasets (axial length > 26 mm), serving as the screening set and the validation set. In the screening set, based on the difference in axial length, 71 SNP loci were identified, among which the key locus with the largest difference was rs62226057.

[0041] For our defined SNP combination, we calculated its AUC values ​​in the validation set at three thresholds: 27 mm, 28 mm, and 29 mm (considered key thresholds for high myopia in the art) to measure the efficacy of this SNP combination in diagnosing high myopia of different severities. Due to the severe imbalance between the randomly assigned case and control samples in the original dataset, we further performed 1:1 random sampling of the case and control groups and recalculated the AUC on the sampled dataset. The results show that the AUC after sampling is highly consistent with the results calculated on the original dataset, indicating robustness of the analysis.

[0042] 5. Statistical analysis: Data are expressed as mean ± standard error.

[0043] II. Experimental Results 1. Participant Demographics: Statistical analysis was performed on the ocular biological parameters of the study participants, with corneal curvature (K) and axial length (AL) as the primary endpoints. After quality control, corneal curvature (K) data from 1341 participants and axial length (AL) data from 1206 participants were obtained. Genome-wide association study (GWAS) was conducted using the 1341 corneal curvature data points as the analysis cohort. Simultaneously, the 1206 axial length data points were randomly divided into a selection set and a validation set at an 11:9 ratio for subsequent locus selection and efficacy evaluation.

[0044] In a GWAS analysis of corneal curvature (K) dimension, a key single nucleotide polymorphism (SNP) associated with corneal curvature, rs62226057 (WNT7B), was identified under significance screening conditions. This locus achieved genome-wide statistical significance in individuals with high myopia (P = 1.974 × 10⁻⁶). -10 This suggests that it may be related to myopia-related changes in ocular biological structures. (See...) Figure 1 ) Because there is a significant negative correlation between corneal curvature and axial length, and the correlation between axial length and high myopia is more direct, the phenotypic effect of this locus was further evaluated in the axial length screening set. The results showed that the rs62226057 locus also had a significant association in the GWAS analysis of axial length (P=6.21×10⁻⁴). After adjusting for covariates such as age and sex, there were significant differences in axial length among samples with different genotypes. The mean axial lengths of individuals with CC, CT, and TT genotypes were 27.32 mm, 27.49 mm, and 27.61 mm, respectively (P=0.0019). This result suggests that the T allele at the rs62226057 locus may be associated with axial elongation and an increased risk of myopia. (See Table 1) Based on this, candidate genetic locus combinations were constructed with rs62226057 as the core, and receiver operating characteristic (ROC) curve analysis was performed on the locus combinations in the axial length screening set and the validation set, respectively. The area under the curve (AUC) was calculated to evaluate its predictive ability for axial elongation and the risk of high myopia, thereby verifying the identification efficacy of the key locus combinations.

[0045] Table 1. Genotype Statistics

[0046] 2. Biomarker screening and verification The basic data of the selected samples are shown in Table 2. A total of 71 SNP combinations were obtained through screening, and their information is shown in Table 3.

[0047] Table 2. Statistics of the Filter Set Data

[0048] Table 3. SNP locus information table

[0049] The efficacy of SNP site combinations in diagnosing high myopia of different severities at different thresholds was verified using a validation set. The validation set sample data is shown in Table 4.

[0050] Table 4. Validation set data statistics

[0051] The AUC obtained for different thresholds in the two datasets are shown in Table 5. Figure 2-3 As shown in Table 3, the SNP locus combinations exhibit good diagnostic efficacy in high myopia of varying severity.

[0052] Table 5. AUC of the original dataset

[0053] 3. Sampling and testing ensure the versatility of the markers. To ensure the identified high myopia risk biomarkers have good generalizability and external validity, a 1:1 random sampling was performed on the case and control groups in the original dataset, and the AUC was recalculated on the sampled dataset. Specifically, an equal number of samples were randomly selected from all HM cases to construct a balanced analysis cohort, avoiding overestimation or bias in model performance due to class imbalance. Both datasets were derived from the same population and constructed using a unified random sampling framework, thereby controlling for confounding factors while maximizing population representativeness. The selected set (Table 6) and validation set (Table 7) data after random sampling are shown in the following tables.

[0054] Table 6. Statistics of the selected set after sampling

[0055] Table 7. Statistics of the Validation Set Data after Sampling

[0056] Refit the dataset on the sampled dataset and calculate the area under the receiver operating characteristic (AUC). The AUCs after sampling are shown in Table 8. Figure 4-5As shown in Table 3, the SNP markers exhibit stable discriminative ability under a balanced sample structure, without significant overfitting. This result not only verifies the reliability of the markers themselves but also demonstrates that the analytical framework constructed through random sampling effectively improves the generalizability of the model in the target population.

[0057] Table 8. AUC of the sampled dataset

[0058] The above description of the embodiments is only for understanding the method and core ideas of the present invention. It should be noted that those skilled in the art can make various improvements and modifications to the present invention without departing from the principles of the invention, and these improvements and modifications will also fall within the protection scope of the claims of the present invention.

Claims

1. A method for constructing a model to assess the risk of high myopia based on axial length, characterized in that, The method includes: obtaining axial length data of highly myopic and non-highly myopic individuals, and constructing a high myopia risk model based on the genotype of SNP markers.

2. The method according to claim 1, characterized in that, The SNP markers are rs62226057: T>C, rs11582005: G>A, rs8191982: G>A, rs199649451: C>T, rs2257107: T>A, rs142759452: C>A, rs6423208: A>G, rs80046353: C>G, rs10932037: C>T, rs3792268: C>A, rs7631705: T>C, rs678690: C>T, rs7507: G>A, rs117234103: G>C, rs31315: T>C, rs77349207: T>C, rs611580: G>C, rs520589: C>T, rs164544: C>T, rs11551053: T>C, rs663227: T>C, rs3823465: A>G, rs2274954: T>C, rs142475387: A>G, rs12678121: C>T, rs80120341: C>T, rs1049832: A>G, rs67210953: G>A, rs4579578: G>T, rs1255411: C>T, rs200502145: G>A, rs56139616: G>A, rs34269909: T>A, rs35607186: T>C, rs34107711: A>G, rs61738000: C>T, rs1356428: A>T, rs76634080: C>G, rs79983883: C>A, rs7138453: T>C, rs59836757: C>T, rs3816503: G>T, rs7306824: G>A, rs79941046: A>G, rs114131205: C>T, rs2075260: G>A, rs2305901: G>A, rs78569455: G>C, rs2876702: T>C, rs3742137: C>T, rs9577407: G>A, rs8017100: T>C, rs2289818: G>C, rs55721315: G>T, rs143952603: G>A, rs8052283: G>A, rs2287078: G>A, rs2967166: A>G, rsrs817330: A combination of C > T, rs12483205: A > G.

3. A kit for assessing the risk of high myopia based on axial length, characterized in that, The kit includes reagents for detecting SNP locus genotypes using one or more methods selected from nucleic acid hybridization, nucleic acid amplification, and sequencing technologies, wherein the SNP locus is the SNP marker described in claim 2.

4. The reagent kit according to claim 3, characterized in that, The reagent is a probe that specifically recognizes the genotype of the SNP site; or, the reagent is a primer that specifically amplifies the gene fragment containing the SNP site.

5. The reagent kit according to claim 3, characterized in that, The kit also includes one or more of the following: reagents for processing samples, standards, calibrators, buffer solutions, and instructions.

6. The use of the reagent for detecting the SNP marker described in claim 2 in a sample in the preparation of products for assessing the risk of high myopia, characterized in that, The products include reagent kits, chips, membrane strips, systems, devices, and storage media.

7. The application according to claim 6, characterized in that, The sample contained nucleic acid from the subject.

8. A system for assessing the risk of high myopia based on axial length, characterized in that, The system includes the following modules: The acquisition module is configured to acquire SNP marker data as described in claim 2 from the sample; The processing module is configured to input the SNP marker data described in claim 2 from the sample into the high myopia risk model constructed by the method described in any one of claims 1-2 for analysis, and obtain the analysis results; The output module is configured to output analysis results.

9. A computer device for assessing the risk of high myopia, characterized in that, The computer device includes a memory and a processor, the memory being used to store computer programs; The processor executes a computer program, which, when executed, implements the following method: Data is acquired to obtain SNP marker data as described in claim 2 from the sample; The data is processed to input the SNP marker data of claim 2 in the sample into the high myopia risk model constructed by the method described in any one of claims 1-2 for analysis, and to obtain the analysis results; Output results, used to output analysis results.

10. A computer program product comprising a computer program for assessing the risk of high myopia, characterized in that, When this computer program is executed by the processor, it implements the following method: Data is acquired to obtain SNP marker data as described in claim 2 from the sample; The data is processed to input the SNP marker data of claim 2 in the sample into the high myopia risk model constructed by the method described in any one of claims 1-2 for analysis, and to obtain the analysis results; Output results, used to output analysis results.

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