Biomarker applied to early diagnosis of sepsis

Abstract

The invention relates to a biomarker applied to early diagnosis of sepsis. The biomarker is a fibrous protein degradation product FDP / D-dimer ratio. Compared with a single index (such as PCT, D-dimer), the biomarker has the advantages that the influence of individual difference and detection error can be reduced, and the stability of a result is improved; in combination with clinical information (such as infection evidence), the ratio can be used as an independent or joint index to assist an existing scoring system (such as SOFA) in improving diagnosis accuracy.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of sepsis diagnosis, and particularly relates to a biomarker applied to early diagnosis of sepsis. BACKGROUND

[0002] Coagulation activation during sepsis is a host response to pathogens and is considered to be one of the causes of tissue damage and multiple organ failure. Hypercoagulability, hypofibrinolysis, microthrombosis and endothelial dysfunction caused by sepsis lead to multiple organ failure. According to statistics, the annual standardized mortality rate of sepsis is 148.1 / 100,000. The detection of conventional coagulation function can indicate the activation of the coagulation system or the systemic inflammatory response, and these indicators can not only indicate the early occurrence of sepsis but also predict the risk of severe inflammation and even death.

[0003] The most important pathophysiological change in sepsis is coagulopathy caused by damage to endothelial cell function and changes in blood flow shear force, which includes an increase in pro-coagulation substances caused by inflammation, such as TF, which is expressed on activated endothelial cells, platelets and mononuclear macrophages in the presence of endotoxin or cytokines (mainly IL-6 and TNF), thereby initiating the coagulation response; sustained consumption of physiological anticoagulants, including APC with anti-inflammatory effects, sustained consumption of antithrombin III, and reduction of TFPI; in addition, the state of first activation and then inhibition of the fibrinolytic system is also found, and studies have shown that endothelial cell activation in the early stage of sepsis leads to an increase in TM concentration, high activation of APC and inhibition of thrombin formation, low activation of TAFI and enhanced fibrinolysis. With the consumption of APC, the generation of thrombin increases, TAFI activation and PAI-1 production increase, and fibrinolysis is inhibited.

[0004] The detection indicators commonly used in current clinical practice include FDP and D-dimer to reflect the fibrinolytic state of sepsis patients. The International Society on Thrombosis and Hemostasis (ISTH) defines the increase of FDP or D-dimer as an important part of the judgment of overt DIC. For the diagnosis of sepsis, the SOFA score is ≥2 points while the infection is simultaneously diagnosed, which is mainly centered on organ failure and cannot achieve early and timely judgment of sepsis, which may delay the treatment of the disease. Endothelial cell damage caused by inflammatory response can be reflected in early changes in coagulation function, which includes common thromboelastography and in vivo anticoagulant system markers, but studies have found that changes in conventional coagulation function occur earlier than TEG. Anticoagulants include early reduction of ATIII, determination of TM, PAI-1 and APC concentration, and considering the economic cost and laboratory conditions, it is not possible to apply them in emergency departments in time and on a large scale.

[0005] Current early diagnosis of sepsis mainly relies on clinical symptoms, inflammatory markers (such as PCT, CRP) and blood culture methods, but these indicators have the following limitations:

[0006] (1)Specificity is insufficient: PCT, CRP and other indicators can be elevated in both infection and non-infectious inflammation, making it difficult to accurately distinguish sepsis from other inflammatory states. For example, PCT may be significantly elevated in patients after severe trauma or major surgery, but does not represent the presence of sepsis, which can lead to misdiagnosis and over-treatment.

[0007] (2) Poor timeliness: Traditional methods such as blood culture take a long time (usually 24-72 hours), and the positive rate is affected by factors such as antibiotic use and sampling time, making it difficult to meet the needs of early diagnosis. During the waiting period for culture results, some patients may have progressed to severe sepsis, delaying the opportunity for early intervention.

[0008] (3) Existing coagulation indicators are single: D-dimer is a marker of coagulation activation and hyperfibrinolysis, and is often elevated in sepsis. However, its use alone has obvious shortcomings: low specificity: D-dimer can also be significantly elevated in non-sepsis conditions such as deep vein thrombosis (DVT), pulmonary embolism (PE), trauma, and malignancy, making it difficult to distinguish sepsis from other hypercoagulable states. High sensitivity but limited discriminative value: Although D-dimer elevation indicates coagulation abnormalities, it cannot directly reflect the pathophysiological processes specific to sepsis (such as endothelial damage and microthrombus formation).

[0009] (4) The dependence and lag of existing scoring systems: SOFA (Sequential Organ Failure Assessment) score and qSOFA (Quick SOFA) score are commonly used tools for sepsis diagnosis, but they mainly rely on clinical symptoms and organ function indicators, which have some subjectivity and may not show obvious abnormalities in the early stages (such as at admission), leading to delayed diagnosis. SUMMARY

[0010] The technical problem to be solved by the present application is to provide a biomarker for early diagnosis of sepsis. Compared with a single indicator (such as PCT, D-dimer), the biomarker can reduce the influence of individual differences and detection errors, and improve the stability of the results.

[0011] The present application provides a biomarker for early diagnosis of sepsis, wherein the biomarker is the ratio of fibrin degradation product FDP to D-dimer D-dimer.

[0012] Further, the biomarker is combined with a pre-set threshold or a machine learning model to determine whether the patient has sepsis.

[0013] Further, the pre-set threshold refers to: if the FDP / D-dimer ratio is ≥2.262, it indicates a high likelihood of sepsis; if the ratio is <2.262, it may be non-sepsis coagulopathy.

[0014] Further, the machine learning model refers to: the input parameters are coagulation indicators and clinical data, and the output result is the probability of sepsis.

[0015] Further, it also includes using the difference between FDP and D-dimer as an auxiliary indicator.

[0016] The application also provides a detection product of a biomarker for early diagnosis of sepsis.

[0017] Further, the detection product includes a kit, test paper or chip.

[0018] Further, the detection uses a patient's venous whole blood.

[0019] The patient refers to a suspected infection patient (meeting the SIRS standard or clinically suspected infection), and the patient's venous whole blood is collected by a citrate sodium-containing anticoagulation tube after admission.

[0020] Further, the patient's venous whole blood is collected for the first time after admission, such as within 6 hours of emergency or ICU admission.

[0021] Further, the detection product is detected by a fully automatic coagulation analyzer.

[0022] Further, the detection method comprises:

[0023] The separated plasma is used for detection, and the detection content includes APTT, PT, TT, Fg, D-dimer and FDP. The blood coagulation instrument is used to detect the concentrations of FDP and D-dimer, and the units are mg / L and mg / L FEU respectively. The detection needs to be completed within 2 hours after blood collection to avoid the influence of sample degradation on the results.

[0024] The calculation formula involved in the application is:

[0025]

[0026] The scientific basis involved in the application is:

[0027] When sepsis occurs, systemic inflammation causes endothelial damage and activation of the coagulation system, and microthrombosis is widely formed and secondary fibrinolysis is enhanced. FDP reflects all fibrin (ogen) degradation products, while D-dimer only represents the degradation fragments of cross-linked fibrin. The microthrombosis of sepsis patients is mainly fibrinogen (non-cross-linked fibrin), so the increase in FDP is often significantly higher than that of D-dimer, resulting in an increase in the ratio; and thrombotic diseases (such as DVT / PE) are mainly cross-linked fibrin, and the increase in D-dimer is more significant, and the ratio is lower.

[0028] Beneficial effects

[0029] 1. Technical effects

[0030] ① Improve diagnostic specificity: Existing sepsis markers (such as PCT, CRP) are easily affected by non-infectious inflammation, while the FDP / D-dimer ratio can better distinguish sepsis from simple thrombotic diseases (such as DVT, PE) or other inflammatory states, reducing the misdiagnosis rate. Through ratio calculation, it can reflect the specific coagulation-inflammation interaction of sepsis (such as microthrombosis and hyperfibrinolysis), which is more relevant to the pathological mechanism than a single indicator (such as D-dimer).

[0031] ② Realize early and rapid screening: Traditional blood culture requires 24-72 hours, while FDP / D-dimer detection only takes 30-60 minutes (based on conventional coagulation analyzers), which can provide auxiliary diagnostic basis when the patient is admitted to the hospital, shortening the clinical decision-making time. It is especially suitable for emergency, ICU and other scenes that need to be quickly judged, helping doctors to start anti-infection treatment in the "golden window period".

[0032] ③ Dynamic monitoring of disease progression: The ratio can be repeatedly detected, and the treatment effect or the severity of sepsis (such as DIC risk) can be evaluated by trend changes (such as retesting at 24 hours and 48 hours after admission).

[0033] 2. Economic effects

[0034] ① Reduce testing costs: FDP and D-dimer are both routine coagulation detection items in clinical practice, without the need for additional equipment or special reagents, and can be completed directly using existing coagulation analyzers, saving additional diagnostic costs. Compared with molecular biology detection (such as gene sequencing) or imaging examination (such as CT), the cost is significantly reduced, making it suitable for promotion in hospitals at all levels.

[0035] ② Reduce waste of medical resources: By accurately identifying sepsis early, the overuse of broad-spectrum antibiotics or ICU resources for non-sepsis patients is avoided, reducing medical expenses.

[0036] 3. Social benefits

[0037] ① Improve patient prognosis: Early diagnosis can intervene in time, reducing the risk of sepsis progressing to severe sepsis, shock or multiple organ failure, and reducing mortality. Through dynamic monitoring of ratio changes, treatment plans (such as the timing of anticoagulant therapy) can be optimized, improving the success rate of treatment.

[0038] ② Improve medical efficiency: Simplify the diagnosis process and reduce the workload of medical staff, especially in areas where medical resources are scarce. ③ Promote the development of precision medicine: Provide new ideas for sepsis subtyping (such as hypercoagulable type vs. hyperfibrinolytic type) and help individualized treatment.

[0039] 4. The comprehensive advantages compared with the prior art

[0040] Comparative dimensions Prior art (e.g. PCT, D-dimer) The present application (FDP / D-dimer ratio) Specificity Low (easily interfered by non-infectious factors) High (based on the coagulation-inflammation interaction mechanism) Timeliness Blood culture takes 24-72 hours Results within 1 hour Detection cost Higher (e.g. PCT detection requires special reagents) Low (using conventional coagulation detection) Pathological relevance Indirect (only reflects a single link of inflammation or coagulation) Direct (reflects the core pathophysiological changes of sepsis)

[0041] 5. Typical application scenarios

[0042] ① Emergency department: rapid screening of patients with fever and coagulation abnormalities, and differentiation of sepsis and common infection.

[0043] ② ICU: dynamic monitoring of coagulopathy progression in patients with sepsis, and guidance of anticoagulant therapy.

[0044] ③ Primary hospital: as an economical and reliable alternative when high-end detection equipment is lacking. BRIEF DESCRIPTION OF DRAWINGS

[0045] Figure 1 The restriction spline curve of the biomarker of the application.

[0046] Figure 2 The nomogram of the logistic regression model of the biomarker of the application. DETAILED DESCRIPTION

[0047] The application will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the application and not to limit the scope of the application. In addition, it should be understood that those skilled in the art can make various modifications or modifications to the application after reading the content taught by the application, and these equivalent forms also fall within the scope defined by the claims attached to the present application.

[0048] Example 1

[0049] 1. Sample collection and processing

[0050] Applicable objects:

[0051] Suspected infected patients (meet at least two of the following) :

[0052] Body temperature > 38℃ or < 36℃;

[0053] Heart rate > 90 times / min;

[0054] Respiratory rate > 20 times / min;

[0055] White blood cell count > 12×10 9 / L or < 4×10 9 / L;

[0056] Clinically suspected infection (such as pneumonia, abdominal infection, urinary tract infection, etc.).

[0057] Blood sampling requirements:

[0058] Blood collection time: the first blood collection after admission (such as emergency triage or within 6 hours of ICU admission).

[0059] Blood collection tube: use sodium citrate anticoagulation tube (blue cap tube, anticoagulation ratio 1:9).

[0060] Sample processing: immediately mix gently for 5-8 times after blood collection, centrifuge at 3000 rpm for 15 minutes at room temperature, separate plasma for detection.

[0061] 2. Detection method

[0062] Detection equipment:

[0063] Fully automatic coagulation analyzer (such as Sysmex CS-2500, STA-R Evolution, ACLTOP 750, etc.).

[0064] Detection items and principles:

[0065] FDP (fibrin degradation product):

[0066] Detection principle: immunoturbidimetry, using anti-FDP antibody to bind with FDP in the sample to form turbidity, and determining the concentration by photometer.

[0067] Unit: mg / L, normal reference range <5 mg / L.

[0068] D-dimer (D-dimer):

[0069] Detection principle: immunofluorescence or ELISA method, specific recognition of cross-linked fibrin degradation products.

[0070] Unit: mg / L FEU (fibrinogen equivalent), normal reference range <0.5 mg / L FEU.

[0071] Quality control:

[0072] Each batch of detection needs to include standard and quality control plasma (such as normal value, high value quality control), to ensure the accuracy of detection.

[0073] 3. Calculate FDP / D-dimer ratio

[0074] Threshold setting (based on clinical research data): as shown in Table 1.

[0075] Table 1. Threshold effect analysis of two linear regression models

[0076]

[0077] Sepsis high risk: ratio ≥2.262 (combined with clinical evidence of infection).

[0078] Low risk or non-sepsis: ratio < 2.262 (e.g. thrombotic disease possible).

[0079] As shown in Figure 1 , a non-linear, inverted U-shaped relationship between FDP / D-dimer ratio (x-axis) and sepsis incidence (y-axis) is shown. The risk peaks at threshold 2.262 (shown by the red dotted line), when the ratio rises to this point, the probability of sepsis will increase.

[0080] 4. Results interpretation and clinical decision

[0081] Sepsis judgment process:

[0082] Step 1: Calculate the ratio and compare it with the preset threshold.

[0083] Step 2: If the ratio is ≥ 2.262 and there is evidence of infection (e.g. positive etiology, imaging of infection focus), start the sepsis process (e.g. antibiotic escalation, fluid resuscitation).

[0084] Step 3: If the ratio is < 2.262 but D-dimer is significantly elevated, rule out thrombotic disease (e.g. DVT, PE).

[0085] Dynamic monitoring application:

[0086] For critically ill patients, the ratio is retested every 24 hours. If the ratio continues to rise, it suggests that sepsis is worsening or there is a risk of DIC.

[0087] 5. Alternative implementation

[0088] Logistic regression model input parameters: FDP, D-dimer, platelet count, PT / APTT and other coagulation indicators, combined with clinical data (APACHE II score, whether receiving anticoagulant therapy, PT value, platelet count and lactic acid concentration).

[0089] Output results: sepsis probability (e.g. Logistic regression model, see Figure 2 ).

[0090] Figure 2This was achieved by visualizing the data graphically, where each influencing factor was assigned a score based on its corresponding value. The total score was then calculated by summing these scores, allowing for the assessment of clinical risk associated with treatment outcomes. For example, an admitted patient had an FDP / D-dimer of 2, an APACHE II score of 30, and was on anticoagulation therapy. Furthermore, their PT was 15 seconds and lactate concentration was 4 mmol / L upon admission. These parameters were assigned scores of 19, 5, 5, 12.5, and 11, respectively. The final calculated total score was 52.5, indicating a low probability of sepsis, corresponding to a predicted sepsis incidence rate of approximately 12%.

Claims

1. A biomarker for early diagnosis of sepsis, characterized in that: The biomarker is the ratio of fibrin degradation product FDP / D-dimer D-dimer.

2. The biomarker according to claim 1, characterized in that: The biomarkers, combined with preset thresholds or machine learning models, determine whether a patient has sepsis.

3. The biomarker according to claim 2, characterized in that: The preset threshold refers to the following: if the FDP / D-dimer ratio is ≥2.262, it indicates a high probability of sepsis; if the ratio is <2.262, it may be a non-septic coagulation abnormality.

4. The biomarker according to claim 2, characterized in that: The machine learning model refers to a model whose input parameters are coagulation indicators and clinical data, and whose output is the probability of sepsis.

5. The biomarker according to claim 1, characterized in that: It also includes using the difference between FDP and D-dimer as an auxiliary indicator.

6. A detection product comprising the biomarkers as described in claim 1 for use in the early diagnosis of sepsis.

7. The testing product according to claim 6, characterized in that: The testing products include reagent kits, test strips, or chips.

8. The testing product according to claim 6, characterized in that: The sample used in the test was the patient's whole venous blood.

9. The testing product according to claim 6, characterized in that: The patient's venous whole blood was collected for the first time after admission.

10. The testing product according to claim 6, characterized in that: The tested products are tested using a fully automated coagulation analyzer.

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