Biomarker combinations and their applications for the auxiliary diagnosis of chronic pain comorbid with depression

Abstract

This invention discloses a combination of biomarkers for the auxiliary diagnosis of chronic pain comorbid with depression and its application, belonging to the fields of in vitro diagnostics (IVD) and clinical testing technology. The biomarker combination includes interleukin-6 (IL-6), interleukin-12 (IL-12), and the concentration ratio of kynurenic acid to 3-hydroxykynurenine (KYNA / 3-HK). This invention also discloses the application of this biomarker combination in an in vitro diagnostic kit for chronic pain comorbid with depression. Experimental results show that the diagnostic efficacy of this biomarker combination (AUC=0.871) is significantly better than that of a single biomarker, effectively distinguishing patients with chronic pain comorbid with depression from those without chronic pain, providing an objective auxiliary diagnostic tool for clinical practice.

Description

Technical Field

[0001] This invention belongs to the fields of in vitro diagnostics (IVD) and clinical testing technology, and specifically relates to a combination of biomarkers for the auxiliary diagnosis of chronic pain comorbid with depression and their application. Background Technology

[0002] In clinical practice, depression is often comorbid with various physical illnesses, with comorbidity with chronic pain being the most common, accounting for approximately 65% ​​of patients with depression. The International Association for the Study of Pain (IASP) defines chronic pain as pain that is persistent or recurrent for more than 3 months. Long-term pain not only reduces patients' quality of life and induces mood disorders, but also impairs social productivity and significantly increases the burden on patients with depression.

[0003] The comorbidity of depression and chronic pain is common, and its diagnosis is a clinical challenge, seriously affecting patient treatment and prognosis. Currently, there is a lack of biological indicators that can objectively and accurately distinguish between depression comorbid with chronic pain (DCP) and simple depression (DNP). Although some studies have explored the roles of inflammatory factors (such as IL-6) or tryptophan metabolites in depression or pain, there are no reports of effective combinations of inflammatory factors and kynurenine metabolites for the auxiliary diagnosis of depression comorbid with chronic pain.

[0004] Therefore, there is an urgent need to develop an objective biomarker-based approach to improve the diagnostic accuracy of chronic pain comorbid with depression. Summary of the Invention

[0005] To address the above deficiencies, this invention provides a combination of biomarkers for the auxiliary diagnosis of chronic pain comorbid with depression. The biomarker combination includes: interleukin-6, interleukin-12, kynurenic acid, and 3-hydroxykynurenine.

[0006] Furthermore, the concentration ratio of kynurenic acid and 3-hydroxykynurenine is used as a combined diagnostic indicator.

[0007] The present invention also includes the use of combining the above-mentioned biomarkers in an in vitro diagnostic kit for the auxiliary diagnosis of chronic pain comorbid with depression.

[0008] Furthermore, the kit includes:

[0009] a) The primary detection reagent for detecting IL-6 concentration in biological samples;

[0010] b) A second detection reagent for detecting the concentration of IL-12 in biological samples;

[0011] c) A third detection reagent for detecting the concentrations of kynurenic acid and 3-hydroxykynurenine in biological samples.

[0012] Furthermore, the auxiliary diagnosis is used to differentiate between patients with depression comorbid with chronic pain and patients with depression alone.

[0013] Furthermore, the biomarker is detected in a biological sample, which may be blood, plasma, or serum.

[0014] Furthermore, the concentrations of interleukin-6 and interleukin-12 were detected using an enzyme-linked immunosorbent assay (ELISA).

[0015] The concentrations of kynurenic acid and 3-hydroxykynurenine were determined by liquid chromatography-tandem mass spectrometry.

[0016] Furthermore, when the concentrations of interleukin-6 and interleukin-12 in the subject's biological sample increased, and the concentration ratio of kynurenic acid to 3-hydroxykynurenine decreased, the subject was identified as a patient with chronic pain comorbid with depression.

[0017] Compared with the prior art, the present invention has the following advantages:

[0018] 1. For the first time, inflammatory factors (IL-6, IL-12) were combined with the kynurenine metabolite ratio (KYNA / 3-HK) to assist in the diagnosis of chronic pain comorbid with depression. The diagnostic efficacy of this combination (AUC=0.871) was significantly better than that of a single biomarker or a combination of some biomarkers.

[0019] 2. The biomarker combination of the present invention can effectively distinguish between patients with depression and chronic pain comorbid with depression and patients with depression without chronic pain, providing an objective auxiliary diagnostic tool for clinical practice.

[0020] 3. The detection methods (ELISA and LC-MS / MS) used in this invention are mature detection technologies that are easy to operate, provide reliable results, and are convenient for clinical application. Attached Figure Description

[0021] Figure 1 This is a bar graph comparing the differences in IL-6 inflammatory factor concentrations in Example 1.

[0022] Figure 2 This is a bar graph comparing the differences in IL-12 inflammatory factor concentrations in Example 1. Detailed Implementation

[0023] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0024] Example 1

[0025] For the study subjects, this embodiment included patients with depression who met the DSM-5 diagnostic criteria, and they were divided into two groups based on whether they had chronic pain:

[0026] Depression with chronic pain group (DCP group): Patients who meet the diagnostic criteria for depression and have chronic pain lasting more than 3 months or intermittent pain for 6 months;

[0027] Depression without chronic pain (DNP group): Patients who meet the diagnostic criteria for depression but do not have chronic pain.

[0028] An age- and sex-matched healthy control group (HC group) was also included.

[0029] Fasting peripheral venous blood was collected from all subjects. After centrifugation, plasma samples were obtained and stored at -80°C for testing. Subsequently, inflammatory factors and kynurenine metabolites were detected.

[0030] 1) Detection of inflammatory factors:

[0031] The concentrations of interleukin-6 (IL-6) and interleukin-12 (IL-12) in plasma were detected by enzyme-linked immunosorbent assay (ELISA). The procedure was performed according to the ELISA kit instructions.

[0032] The experimental results are shown in Table 1 below:

[0033] Table 1

[0034]

[0035] In Table 1 above, H is the Kruskal-Walis test statistic, F is the one-way ANOVA statistic, and P-adj is the p-value after post-hoc correction.

[0036] Bar graph comparing differences in inflammatory factor concentrations, as shown below Figure 1-2 As shown, * represents P0.05 and ** represents P0.001.

[0037] 2) Detection of kynurenine metabolites:

[0038] The concentrations of kynurenic acid (KYNA) and 3-hydroxykynurenic acid (3-HK) in plasma were determined by liquid chromatography-tandem mass spectrometry (LC-MS / MS), and the concentration ratio of KYNA to 3-HK (KYNA / 3-HK) was calculated.

[0039] The experimental results are shown in Table 2 below:

[0040] Table 2

[0041]

[0042] In the table above, H is the Kruskal-Wallis test statistic, F is the one-way ANOVA statistic, and P-adj is the p-value after post-hoc correction.

[0043] Therefore, the following comparison can be made:

[0044] Comparison of inflammatory factor levels:

[0045] The IL-6 concentration in the DCP group was 86.856 (117.578) pg / ml, which was significantly higher than that in the DNP group (62.820 ± 33.494 pg / ml) and the HC group (P<0.05).

[0046] The IL-12 concentration in the DCP group was 5.428 (2.183) pg / ml, which was significantly higher than that in the DNP group (4.614 ± 1.767 pg / ml) and the HC group (P<0.05).

[0047] Comparison of canine urinary tract metabolite ratios:

[0048] The KYNA / 3-HK ratio in the DCP group was 0.384 (0.235), which was significantly lower than that in the DNP group (0.565 (0.395)) and the HC group (P<0.001), suggesting that the kynurenine metabolic pathway in DCP patients is shifted towards neurotoxicity.

[0049] The combined effect was verified, as shown in Table 3 below:

[0050] Table 3

[0051]

[0052]

[0053] As shown in the table above, while both the combination of inflammatory pathway biomarkers and the single use of tryptophan metabolism pathway biomarkers have some diagnostic value (AUCs of 0.701 and 0.692, respectively), their efficacy is limited. However, when the three biomarkers defined in this invention—IL-6, IL-12, and KYNA / 3-HK—from two different pathways are used in combination, the AUC value of the combined diagnostic model is significantly increased to 0.823. This result clearly confirms that the biomarker combination proposed in this invention has an unexpected synergistic effect, and its diagnostic efficacy is significantly better than any single pathway biomarker combination.

[0054] Example 2

[0055] This embodiment describes the application of the biomarkers from Example 1 in the preparation of an in vitro diagnostic kit for assisting in the diagnosis of patients with depression comorbid with chronic pain and patients with simple depression.

[0056] The kit includes:

[0057] a) The first detection reagent used to detect the concentration of IL-6 in biological samples (detected by enzyme-linked immunosorbent assay).

[0058] b) A second detection reagent for detecting the concentration of IL-12 in biological samples (using enzyme-linked immunosorbent assay).

[0059] c) A third detection reagent for detecting the concentrations of kynurenic acid and 3-hydroxykynurenine in biological samples (using liquid chromatography-tandem mass spectrometry).

[0060] Biomarkers are detected in biological samples, such as blood, plasma, or serum.

[0061] When the concentrations of interleukin-6 and interleukin-12 in a subject's biological sample are elevated, and the concentration ratio of kynurenic acid to 3-hydroxykynurenine is decreased, the subject is identified as a patient with chronic pain comorbid with depression.

[0062] In detail, this kit is used to detect the levels of the three biomarkers mentioned above in a biological sample (preferably plasma) of an individual being tested. The kit may contain the necessary buffer solution, standards, ELISA plate, and detection antibodies. The kit may also include instructions that contain reference values, cut-off values, or recommended calculation models / formulas for indicating the risk of chronic pain comorbid with depression, to help users interpret the test results.

[0063] The practical steps are as follows:

[0064] S1. Obtain biological samples from the individual to be tested;

[0065] S2. Detect the concentrations of IL-6, IL-12, KYNA, and 3-HK in the sample;

[0066] S3. Calculate the KYNA / 3-HK ratio;

[0067] S4. Compare the test results with a preset reference range or threshold, or substitute the results into a diagnostic model to obtain the risk assessment results of the individual having depression comorbid with chronic pain.

[0068] It should be noted that the structure described in this invention can be implemented in many different forms and is not limited to the embodiments described. Any equivalent transformations made by those skilled in the art based on the content of this specification, or direct or indirect applications in other related technical fields, are included within the protection scope of this invention.

Claims

1. A combination of biomarkers for the auxiliary diagnosis of chronic pain comorbid with depression, characterized in that, The biomarker combination includes: interleukin-6, interleukin-12, kynurenic acid, and 3-hydroxykynurenine.

2. The biomarker combination as described in claim 1, characterized in that: The concentration ratio of kynurenic acid and 3-hydroxykynurenine was used as a combined diagnostic indicator.

3. The use of the biomarker described in claim 1 in an in vitro diagnostic kit for the auxiliary diagnosis of chronic pain comorbid with depression.

4. The application as described in claim 3, characterized in that: The kit includes: a) The primary detection reagent for detecting IL-6 concentration in biological samples; b) A second detection reagent for detecting the concentration of IL-12 in biological samples; c) A third detection reagent for detecting the concentrations of kynurenic acid and 3-hydroxykynurenine in biological samples.

5. The application as described in claim 3, characterized in that: The auxiliary diagnosis is used to differentiate between patients with depression and chronic pain comorbid with depression and patients with depression alone.

6. The application as described in claim 3, characterized in that: The biomarker is detected in a biological sample, which may be blood, plasma, or serum.

7. The application as described in claim 3, characterized in that: The concentrations of interleukin-6 and interleukin-12 were detected by enzyme-linked immunosorbent assay (ELISA). The concentrations of kynurenic acid and 3-hydroxykynurenine were determined by liquid chromatography-tandem mass spectrometry.

8. The application as described in claim 3, characterized in that: When the concentrations of interleukin-6 and interleukin-12 in a subject's biological sample are elevated, and the ratio of kynurenic acid to 3-hydroxykynurenine concentration is decreased, the subject is identified as a patient with chronic pain comorbid with depression.

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