Use of cathepsin h in peripheral blood serum in diagnosis and treatment of alzheimer's disease

Abstract

This invention relates to the use of cathepsin H in peripheral blood serum in the diagnosis and treatment of Alzheimer's disease, belonging to the field of biotechnology. Through research and verification of the expression / activity characteristics of CatH in AD-related samples and its relationship with disease development, this invention establishes serum CatH as a novel diagnostic and therapeutic target / biomarker for AD, enabling peripheral fluid detection for screening and auxiliary diagnosis. It also provides a basis for screening inhibitors targeting CatH and developing peripheral intervention drugs.

Description

Technical Field

[0001] This invention relates to the use of cathepsin H in peripheral blood serum in the diagnosis and treatment of Alzheimer's disease, and belongs to the field of biotechnology. Background Technology

[0002] Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive cognitive decline and reduced daily living abilities. It is characterized by insidious onset, prolonged course, and high disability rate. With the increasing aging of the population, the number of AD patients continues to rise, leading to a growing clinical need for early identification, pathological confirmation, treatment efficacy evaluation, and long-term management.

[0003] Currently, the biological assessment of Alzheimer's disease (AD) primarily relies on molecular imaging and humoral biomarker detection. Clinical research and some clinical practices often employ the A / T / (N) framework to characterize amyloid-β (Aβ) deposition (A), pathological tau (T), and neuronal damage / degeneration (N), thereby aiding in pathological confirmation, classification, and disease progression assessment. Commonly used methods include molecular imaging examinations such as Aβ-PET, tau-PET, and FDG-PET, as well as the detection of Aβ and tau in cerebrospinal fluid (CSF). In recent years, blood biomarkers have become an important research direction for the early identification and large-scale diagnosis of AD due to their convenient sampling, relatively low cost, repeatable monitoring, and suitability for screening.

[0004] In terms of treatment, current AD intervention strategies mainly include symptomatic treatment and disease-modifying therapy (DMT). Symptomatic drugs, such as the NMDA receptor antagonist memantine, are used to improve symptoms in some patients. Some progress has been made in the field of DMT, with representative strategies including monoclonal antibody therapy targeting Aβ, which is suitable for specific populations and emphasizes assessment of its impact on disease progression. Meanwhile, the high selectivity of the blood-brain barrier (BBB) ​​poses challenges to central drug administration and effective drug exposure, driving research into alternative or complementary pathways from the perspectives of translatability and clinical accessibility.

[0005] However, existing technologies still have the following shortcomings: (I) Diagnostic deficiencies: 1. Limited accessibility of PET molecular imaging examinations: PET molecular imaging examinations are costly and require high levels of equipment, tracer supply, and image interpretation capabilities, making it difficult to achieve large-scale application in grassroots or resource-limited areas. 2. Invasiveness and acceptability issues with CSF biomarker detection: Collection relies on lumbar puncture, limiting its promotion in routine outpatient clinics and large-scale screening scenarios. 3. The specificity and interpretability of blood biomarkers are affected by peripheral confounding factors: In the context of multiple coexisting diseases and complex medication use in the elderly, factors such as peripheral inflammatory status, changes in liver and kidney function, and cardiovascular and metabolic abnormalities may cause fluctuations in levels, reducing their specificity for the core pathology of AD and the interpretability of results. 4. Stable, reliable blood biomarkers with clear association with AD pathology still need further screening and validation: There are still technical gaps in improving diagnostic coverage, accuracy, stratified management, and efficacy monitoring. (II) Treatment-Related Deficiencies: 1. Symptomatic treatment is unlikely to alter the natural course of the disease: Existing symptomatic drugs are mostly used to improve or stabilize some symptoms, but they are usually unable to fundamentally block or reverse the neurodegenerative process, and the benefits vary from person to person. 2. Existing DMTs have limitations in the applicable population and the extent of benefits: Strategies represented by anti-Aβ monoclonal antibodies are generally applicable to people in the early stages and with evidence of Aβ pathology. Clinical benefits are mostly reflected in a slower rate of decline, and the scope of application is relatively limited. 3. The blood-brain barrier limits effective exposure to central nervous system drugs: The high selectivity of the blood-brain barrier makes it difficult for many candidate drugs to enter the central nervous system and achieve effective exposure, increasing the complexity of research and development in terms of route of administration selection, dosage optimization, and safety balance. 4. There is still a lack of new targets and strategies with greater translational potential and controllable risks: In terms of efficacy, safety, accessibility, and long-term management, it is still necessary to further clarify the key pathways related to disease progression and propose new intervention pathways.

[0006] Given the current situation, the industry is increasingly focusing on molecular targets that are detectable in the peripheral circulation, relatively stable, and druggable, aiming to achieve earlier assessment and intervention of AD pathological processes while reducing invasiveness and improving accessibility. If a target closely related to the occurrence and development of AD can be identified and stably detected in the circulation, and a technical pathway can be built around this target, from target validation to candidate inhibitor screening and evaluation, especially through computational methods such as molecular docking to assist in the discovery of potential inhibitors and the research and application of peripheral drug delivery interventions, it is expected to provide a new approach to the integrated diagnosis and treatment of AD and lay a technical foundation for subsequent drug development and translational applications. Summary of the Invention

[0007] In view of this, the object of the present invention is to provide the use of cathepsin H in peripheral blood serum in the diagnosis and treatment of Alzheimer's disease.

[0008] To achieve the above objectives, the technical solution of the present invention is as follows.

[0009] The use of peripheral blood cathepsin H as a biomarker in the preparation of products for the diagnosis and / or auxiliary diagnosis of Alzheimer's disease.

[0010] The use of peripheral blood cathepsin H as a target in the preparation of products for the treatment of Alzheimer's disease.

[0011] Use of reagents for determining cathepsin H levels in peripheral blood in the preparation of diagnostic kits for Alzheimer's disease.

[0012] The use of cathepsin H inhibitors in the preparation of drugs for treating Alzheimer's disease.

[0013] Furthermore, the cathepsin H inhibitor is the CatH inhibitor E64.

[0014] Furthermore, the administration method is peripheral administration.

[0015] Furthermore, cathepsin H inhibitors reduce inflammation-related responses in the brain;

[0016] And / or, cathepsin H inhibitors reduce Aβ deposition; And / or, cathepsin H inhibitors improve BBB permeability.

[0017] Beneficial effects This invention establishes CatH in serum as a new diagnostic and therapeutic target / biomarker for AD by studying and verifying the expression / activity characteristics of CatH in AD-related samples and its relationship with the occurrence and development of the disease. Based on this, peripheral body fluid detection can be used for screening and auxiliary diagnosis, and at the same time, it provides a basis for the screening of CatH inhibitors and the development of peripheral intervention drugs. Attached Figure Description

[0018] Figure 1 This is a diagram showing the localization of CatH in brain vascular endothelial cells under AD background in Example 1, and the characteristics of its expression and polarized secretion induced by Aβ.

[0019] Figure 2 The graph shows the cognitive impairment level and serum CatH level of 7-month-old WT and AD mice in Example 2.

[0020] Figure 3 The results of the test in Example 3 showed that peripheral administration of the CatH inhibitor significantly improved the cognitive ability of AD mice.

[0021] Figure 4 This is the test result related to the significant reduction in AD-like pathological burden in the brain when peripheral CatH inhibition was administered in Example 4. Detailed Implementation

[0022] The present invention will be further described in detail below with reference to specific embodiments.

[0023] This invention provides the use of peripheral blood cathepsin H as a biomarker in the preparation of products for the diagnosis and / or auxiliary diagnosis of Alzheimer's disease.

[0024] This invention provides the use of peripheral blood cathepsin H as a target in the preparation of products for treating Alzheimer's disease.

[0025] This invention provides the use of reagents for determining cathepsin H levels in peripheral blood in the preparation of diagnostic kits for Alzheimer's disease. Examples include specific binding reagents for detecting cathepsin H protein levels, and / or substrates / probes for detecting cathepsin H activity levels.

[0026] This invention provides the use of cathepsin H inhibitors in the preparation of drugs for treating Alzheimer's disease.

[0027] Furthermore, the cathepsin H inhibitor is the CatH inhibitor E64.

[0028] Furthermore, the administration method is peripheral administration.

[0029] Furthermore, cathepsin H inhibitors reduce inflammation-related responses in the brain; And / or, cathepsin H inhibitors reduce Aβ deposition; And / or, cathepsin H inhibitors improve BBB permeability.

[0030] An in vitro detection method for Alzheimer's disease, characterized by comprising the following steps: a) Obtain peripheral biological samples from the subjects; b) Detect the expression level and / or activity level of cathepsin H in the peripheral biological sample to obtain the detection value; c) Compare the detected value with the reference value; d) When the detected value increases relative to the reference value and / or meets the preset judgment criteria, the subject is determined to have an increased risk of Alzheimer's disease, and / or is an Alzheimer's disease patient, and / or is used to assess the disease stage / severity of Alzheimer's disease.

[0031] A pharmaceutical composition for the prevention and / or treatment of Alzheimer's disease, characterized in that the pharmaceutical composition comprises any of the following active ingredients: a) a cathepsin H inhibitor; b) an antibody against cathepsin H or an antigen-binding fragment thereof; c) a vector and / or viral vector expressing a nucleic acid molecule for inhibiting cathepsin H expression; wherein the pharmaceutical composition is capable of reducing the activity and / or expression level of cathepsin H.

[0032] A method for preparing the pharmaceutical composition, characterized by comprising the following steps: S1: Obtain candidate inhibitors of cathepsin H and perform activity screening and confirmation; S2: Prepare a pharmaceutical formulation by combining the identified inhibitor with a pharmaceutically acceptable carrier and / or excipient; S3: Prepare antibody / antigen-binding fragments against cathepsin H; and / or construct nucleic acid molecules and their delivery vectors / viral vectors for inhibiting cathepsin H expression.

[0033] This invention proposes and establishes catepsin H (CatH) as a novel diagnostic and therapeutic target / blood biomarker for Alzheimer's disease (AD). Quantitative detection of CatH activity in peripheral serum is used for screening, auxiliary diagnosis, and assessment of disease course / severity (including efficacy monitoring). A transferable peripheral intervention strategy is constructed: by inhibiting CatH activity / expression through peripheral drug administration (inhibitors, antibodies, or nucleic acids, etc.), improvements in AD-related pathology and cognitive phenotypes can be achieved.

[0034] Example 1: Localization of CatH in brain vascular endothelial cells and Aβ-induced polarization secretion characteristics in the context of AD. (1) Observation of immunofluorescence co-staining and co-localization of brain tissue Seven-month-old 5×FAD mice were selected and anesthetized with isoflurane inhalation followed by 4% paraformaldehyde (PFA) perfusion fixation. Brain tissue was then removed, post-fixed, and dehydrated. Brain tissue sections approximately 30 μm thick were prepared using frozen sectioning. The sections were then subjected to double immunofluorescence staining using the bleaching method, with CatH as the detection target and CD31 as a vascular / endothelial cell marker. After staining, the sections were imaged and analyzed using laser confocal microscopy. Imaging results showed that CatH and CD31 signals significantly overlapped in the vascular structure region. Figure 1 A) suggests that CatH has localization characteristics in brain vascular endothelial cell-related regions.

[0035] (2) In vitro verification of the expression and polarization secretion characteristics of CatH in brain endothelial cells under Aβ stimulation To investigate the possible driving factors of increased CatH expression in brain vascular endothelial cells under AD background, the mouse brain microvascular endothelial cell line b.End3 was used as an in vitro model to simulate Aβ-related stimulation and detect changes in CatH expression. Specifically, b.End3 cells were treated with 2 μM Aβ, and cell samples were collected at 12 h, 24 h, and 48 h after stimulation; the control group was treated with an equal volume of solvent (DMSO). Total RNA was extracted from the cells, and the expression level of CatH mRNA was detected by quantitative real-time PCR (qPCR). The results showed that compared with the control group, Aβ stimulation increased the expression level of CatH mRNA in b.End3 cells, and the expression level increased with prolonged stimulation time. Figure 1 B).

[0036] Based on this, to further analyze the secretion characteristics of CatH, an in vitro blood-brain barrier model was constructed using the Transwell system. A dense monolayer of b.End3 cells was cultured on a polycarbonate membrane. The upper chamber was defined as the blood side (apical / cavitary side), and the lower chamber as the brain parenchyma side (basolateral / epidermal side). Aβ stimulation was applied to the brain parenchyma side, and the culture supernatants from both chambers were collected and CatH levels were measured. The results showed that under Aβ stimulation on the brain parenchyma side, the CatH level in the blood side supernatant increased (…). Figure 1 C, D).

[0037] To further analyze the spatial distribution changes of CatH within cells, cells on the Transwell membrane were stained with CatH immunofluorescence and analyzed using Z-axis tomography with laser confocal microscopy. The distribution of CatH signal along the Z-axis was compared between the control group and the Aβ stimulation group under the condition of consistent tomographic range. Imaging results showed that the CatH signal in the control group was relatively uniformly distributed along the Z-axis; however, after 48 h of Aβ stimulation on the brain parenchyma side, the CatH signal shifted towards the blood side along the Z-axis. Figure 1 E and F) suggest that CatH exhibits polarized distribution changes in endothelial cells.

[0038] In summary, under the conditions of this implementation, CatH has localization characteristics in brain vascular endothelial cells under AD background, and Aβ stimulation on the brain parenchyma side can induce upregulation of CatH expression in endothelial cells, accompanied by changes in its secretion and distribution to the blood side.

[0039] Figure 1This study demonstrates the localization of CatH in brain vascular endothelial cells under AD background and the characteristics of its expression and polarized secretion induced by Aβ. A, Immunofluorescence staining of brain slices from 7-month-old 5xFAD mice, showing significant co-localization of the vascular endothelial cell marker CD31 with CatH; B, CatH mRNA levels in b.End3 cells 12 h, 24 h, and 48 h after Aβ stimulation; C, CatH levels in the blood-side culture medium after Aβ stimulation on the brain parenchyma side; D, Gray value analysis of CatH protein levels in the blood-side culture medium in Figure C; E, Distribution of CatH signal on the Z-axis in cells after Aβ stimulation on the brain parenchyma side; F, Quantitative analysis of CatH fluorescence signal in Figure E. Scale bar: 50 µm. Two-tailed Student's t-test; *p<0.05, **p<0.01, ***p<0.001.

[0040] Example 2: Cognitive impairment in 7-month-old WT and AD mice was positively correlated with serum CatH levels. (1) Serum sample collection and CatH quantitative detection Seven-month-old WT mice and 5×FAD mice were selected. Peripheral blood was collected and serum samples were obtained. Serum CatH levels were quantitatively measured using an ELISA kit targeting catepsin H (CatH). The results showed that compared with age-matched WT mice, the serum CatH level in 7-month-old 5×FAD mice was significantly higher. Figure 2 A).

[0041] (2) Cognitive function behavioral assessment To assess the cognitive functions of the test mice, such as learning and working memory, behavioral tests including the Morris Water Maze (MWM), Novel Object Recognition (NOR), and Y-maze were used.

[0042] Morris Water Maze (MWM): The experiment lasted for 7 days. For the first 5 days, mice underwent orienteering training, four times a day, starting from different quadrant entry points. The time required for the mice to reach the hidden platform within a specified time (latency period) was recorded. On the 7th day, a spatial exploration experiment was conducted. The platform was removed, and the mice were placed in the water tank from designated quadrant entry points. Their swimming trajectories within a specified time were recorded, and spatial memory indicators such as the percentage of the path to the target quadrant were calculated.

[0043] Novelty Object Recognition (NOR): In the familiarization phase, two identical objects are placed in a designated location within a test chamber, and the exploration time of the mouse for each object is recorded within a limited time. After a certain interval, in the testing phase, one of the objects is replaced with a new object, and the exploration time of the mouse for the new object and the old object is recorded. The recognition index (RI) is calculated using the following formula: RI = (New object exploration time / (New object exploration time + Old object exploration time)) × 100%.

[0044] Y-maze: Mice are placed in the central area of ​​the maze and allowed to explore freely for a predetermined time. The order and number of arm entries are recorded, and the spontaneous alternation rate is calculated based on the number of "alternations" formed by consecutive entries into three different arms. The spontaneous alternation rate is calculated using the following formula: Spontaneous alternation rate (%) = [Number of spontaneous alternations / (Total number of arm advances)] 2) × 100%.

[0045] Behavioral test results showed that, compared with WT mice of the same age, 7-month-old 5×FAD mice exhibited a trend of decreased cognitive function in the above-mentioned cognitive-related indicators. Figure 2 BD).

[0046] (3) Correlation analysis between serum CatH and cognitive function indicators Linear correlation analysis was performed on the serum CatH quantification results of each mouse and their behavioral test indicators. The results showed that serum CatH levels were correlated with the degree of cognitive impairment; that is, when serum CatH levels increased, the cognitive performance of mice decreased accordingly. Figure 2 (EG). The above results indicate that serum CatH levels can be used to reflect cognitive impairment-related changes in AD model mice, suggesting its potential as a peripheral blood biomarker for assessing AD-related pathological processes, determining staging / severity, and monitoring treatment efficacy.

[0047] Figure 2This study demonstrates a positive correlation between cognitive impairment and serum CatH levels in 7-month-old WT and AD mice. A, Serum CatH levels in 7-month-old WT and 5×FAD mice; B, Spontaneous alternation rate in the Y-maze test in 7-month-old WT and 5×FAD mice; C, Cognitive index of mice in the novelty recognition test; D, Time to reach the platform in the water maze test; E, Correlation analysis between cognitive performance and serum CatH levels in 7-month-old WT and 5×FAD mice in the Y-maze (E), novelty recognition (F), and water maze (G). Two-way ANOVA or two-tailed Student's t-test, *p<0.05, **p<0.01, ***p<0.001.

[0048] Example 3: Peripheral administration of CatH inhibitors significantly improved cognitive abilities in AD mice To evaluate the intervention effect of peripheral catepsin H (CatH) inhibitor on Alzheimer's disease (AD)-related phenotypes, 4-month-old wild-type (WT) mice and 5×FAD transgenic mice were selected for drug treatment. The experimental group was administered the CatH inhibitor E64 intraperitoneally at a dose of 15 mg / kg, once every 3 days for 2 months; the control group received an equal volume of PBS (PBS) with the same frequency and volume of administration. Figure 3 A). After drug administration, the mice were behaviorally assessed to evaluate the feasibility of CatH as a potential therapeutic target.

[0049] Behavioral assessments include the open field test (OFT), novel object recognition (NOR), and Y-maze, which are used to assess general activity level / anxiety-like behavior, learning and memory abilities, and spatial working memory, respectively.

[0050] Open field test (OFT): Mice were placed in the test room for at least 24 hours to acclimatize to the environment before the test. The mice were placed in the central area of ​​the open field device and allowed to explore freely for 10 minutes. The movement trajectory was recorded by a behavioral data acquisition system, and the time spent in the central area and the total amount of movement were statistically analyzed.

[0051] Novelty Object Recognition Experiment (NOR): Conducted according to the method described in Example 2, including a familiarization phase and a testing phase, recording the exploration time for new and old objects and calculating the recognition index (RI).

[0052] Y-maze experiment: The experiment was conducted as described in Example 2. The order of arm entry was recorded and indicators such as the spontaneous alternation rate were calculated.

[0053] The results showed that in the open field experiment, after 2 months of peripheral administration of E64, the central region dwell time and other indicators related to anxiety-like behavior in WT mice and 5×FAD mice did not show a trend indicating adverse changes, while the overall activity / exploration level showed an upward trend. Figure 3 B). In the novelty recognition test and the Y-maze test, compared with the PBS control group, E64-treated 5×FAD mice showed improvements in learning memory and spatial working memory related indicators. Figure 3 (CE). The above results indicate that, under the conditions described in this study, inhibiting CatH activity via peripheral administration can improve cognitive-related behavioral performance in 5×FAD mice, suggesting that peripheral inhibition of CatH has the potential for application in AD intervention.

[0054] Figure 3 This study demonstrates that peripheral administration of CatH inhibitors significantly improved cognitive abilities in AD mice. A, Schematic diagram of the E64 intervention experiment; B, Time spent in the central region by mice in the open field test; C, Cognitive index of mice in the novelty recognition test; D, Heatmap of mouse movement trajectories in the novelty recognition test; E, Spontaneous alternation rate of mice in the Y-maze test. Two-way ANOVA; *p<0.05, **p<0.01, ***p<0.001.

[0055] Example 4: Peripheral administration of CatH significantly alleviated the AD-like pathological burden in the brain. After the dosing regimen described in Example 3 was completed, in order to further evaluate the effect of peripheral inhibition of CatH activity on AD-related intracranial pathological changes and blood-brain barrier function, brain tissue samples were collected and tested from WT mice and 5×FAD mice that received intraperitoneal injections of PBS or E64.

[0056] (1) Immunofluorescence detection of microglia activation and Aβ plaque burden in brain tissue sections Brain tissue sections were prepared from mice in each group and subjected to immunofluorescence staining. The microglia marker Iba1 was used to assess microglial activation levels, and the Aβ-related antibody 6E10 was used to assess Aβ plaque burden. Fluorescence microscopy and image analysis showed that, compared with the PBS control group, E64-treated 5×FAD mice exhibited decreased microglial activation-related signals and reduced Aβ plaque burden. Figure 4 A). The above results suggest that, under the conditions described in this study, peripheral E64 intervention was associated with reduced inflammation-related responses and decreased Aβ deposition in the brains of 5×FAD mice.

[0057] (2) Detection of Aβ1-42 and inflammation-related factors in brain tissue Further analysis of brain tissue homogenate samples was conducted to detect the expression levels of Aβ1-42 and inflammation-related factors (using methods such as ELISA, qPCR, or Western blotting, depending on the specific conditions). Results showed that compared to the 5×FAD group treated with PBS, the E64-treated group exhibited decreased Aβ1-42 levels and a reduction in the expression levels of multiple inflammation-related factors. Figure 4 B. Figure 4 E). This result is consistent with histological observations.

[0058] (3) Evans blue tracer to assess blood-brain barrier permeability To determine whether peripheral inhibition of CatH activity could improve barrier damage caused by increased BBB permeability, an Evans blue tracer assay was used to assess BBB permeability. The specific steps were as follows: 50 µL of 60 mg / mL Evans blue solution was injected into mice via the tail vein; one hour after injection, the mice were perfused with PBS to remove residual dye from the blood vessels, and then the brain was harvested. The cortical tissue was separated, weighed, and placed in a 1.5 mL centrifuge tube. PBS was added at a ratio of 500 µL per 100 mg of brain tissue, and the mixture was thoroughly homogenized. Then, an equal volume of formamide was added, and the mixture was incubated at 55 °C for 24 hours to extract the dye. After incubation, the mixture was centrifuged at 1000 rpm for 5 minutes, and the supernatant was collected. The absorbance of the supernatant at 620 nm and 740 nm was measured using a microplate reader, and the result was expressed as "(OD620)". "OD740 / brain tissue or weight" is used as a quantitative indicator of Evans blue extravasation to characterize the degree of BBB damage.

[0059] Experimental observations showed that visible blue staining appeared in the brain tissue of the 5×FAD group treated with PBS, indicating increased BBB permeability; in contrast, the degree of blue staining in the brain tissue of the 5×FAD group treated with E64 was reduced. Figure 4 C). Further quantitative results showed a difference between the E64 treatment group and the PBS control group ( Figure 4 D).

[0060] Figure 4 This study demonstrates that peripheral administration of CatH significantly alleviates the burden of AD-like pathological conditions in the brain. A, Immunofluorescence staining of brain slices from WT and 5×FAD mice after intraperitoneal injection of PBS / E64, including Hoechst nuclear dye and the microglial marker Iba1; B, ELISA detection of Aβ in mouse brain tissue. 1–42C, Quantitative analysis of Evans blue dye in mouse brain tissue; D, Mouse brain tissue after Evans blue injection; E, Heatmap of inflammatory factor expression levels in mouse brain tissue. Two-tailed Student's t-test; *p<0.05, **p<0.01, ***p<0.001.

[0061] In summary, under the conditions described in this study, peripheral administration of E64 to inhibit CatH activity was associated with reduced inflammation-related responses, decreased Aβ deposition, and improved BBB permeability in the brains of 5×FAD mice. Combined with the behavioral results in Example 3, this suggests that peripheral inhibition of CatH has the potential for application in AD intervention.

[0062] This invention uses serum to detect the protein and / or activity levels of CatH. Peripheral blood collection is relatively simple, allows for repeated dynamic monitoring, and is naturally suitable for outpatient screening and follow-up. CatH, as a "detectable protease on the circulating side," avoids the high-barrier path of relying on PET / CSF to obtain pathological information. This invention enhances the pathological relevance of the indicator by validating evidence of the brain origin / localization of cathepsin H, combined with peripheral detection evidence and behavioral phenotype-related evidence, avoiding the blood biomarker remaining merely at the level of "statistical difference." In particular, the association between endothelial-related localization and BBB damage provides a more interpretable bridge for "how peripheral level changes reflect central pathological progression." It provides a peripheral intervention pathway targeting CatH. This strategy does not rely on high-cost imaging typing as the sole prerequisite, nor does it require a complete adoption of the "anti-Aβ monoclonal antibody infusion + imaging monitoring" model. Targeting a protease provides a basis for drugability, and the peripheral administration route makes it easier to optimize dosage form and dosing frequency. Peripheral administration can directly act on the circulating target without relying on drugs to achieve high exposure across the BBB.

[0063] In summary, the invention includes, but is not limited to, the above embodiments. Any equivalent substitutions or partial improvements made under the spirit and principles of this invention shall be considered to be within the protection scope of this invention.

Claims

1. Use of peripheral blood cathepsin H as a biomarker in the preparation of products for the diagnosis and / or auxiliary diagnosis of Alzheimer's disease.

2. The use of peripheral blood cathepsin H as a target in the preparation of products for the treatment of Alzheimer's disease.

3. Use of reagents for determining the level of cathepsin H in peripheral blood in the preparation of diagnostic kits for Alzheimer's disease.

4. The use of cathepsin H inhibitors in the preparation of drugs for treating Alzheimer's disease.

5. The use as described in claim 4, characterized in that: The cathepsin H inhibitor is CatH inhibitor E64.

6. The use as described in claim 4, characterized in that: The administration route is peripheral.

7. The use as described in claim 4, characterized in that: Inhibitors of cathepsin H reduce inflammation-related responses in the brain; And / or, cathepsin H inhibitors reduce Aβ deposition; And / or, cathepsin H inhibitors improve BBB permeability.

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