Application of H1152 in preparation of medicine for treating metabolism-related cognitive impairment

Abstract

The invention discloses an application of H1152 in preparation of a medicine for treating metabolism-related cognitive impairment, belongs to the technical field of medicine, breaks through the limitation that an existing medicine singly regulates metabolism or cognition, and the H1152 can synchronously inhibit abnormal weight growth induced by high fat diet and remarkably improve spatial memory water maze experiments and short-term memory new object recognition experiments. Dual intervention of metabolism and nerves is realized; compared with statins, the H1152 does not observe creatine kinase rise or liver enzyme abnormity under the equivalent lipid-lowering dosage, and animal experiments show that the H1152 has better safety; the ROCK signal channel is regulated and controlled, and the ROCK signal channel acts on vascular endothelial function improvement and neuronal synaptic plasticity enhancement at the same time, so that the problem of co-disease of metabolism and cognition is solved from the multi-target level; the body weight growth rate, the water maze escape latency and the new object recognition index are integrated as the combined curative effect markers for the first time, and quantifiable objective standards are provided for clinical curative effect evaluation.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technologies, and particularly relates to the use of H1152 in the preparation of a medicament for treating metabolic-related cognitive impairment. Background Art

[0002] In the prior art, APOE gene knockout animal models have been proven to simulate the pathological process of human metabolic syndrome combined with cognitive decline. Although statins commonly used clinically can improve blood lipid metabolism currently, their protective effect on cognitive function is limited, and there are side effects such as myotoxicity. As a ROCK inhibitor, previous studies on H1152 have mainly focused on the field of cardiovascular diseases, and there has been no report on its combined intervention in metabolic-cognitive comorbidities.

[0003] Application of APOE gene knockout animal models in the study of metabolic-cognitive comorbidities, model construction and pathological characteristics: APOE gene knockout (ApoE KO) animals (such as mice, dogs) are achieved through CRISPR / Cas9 technology and can spontaneously develop hypercholesterolemia and atherosclerosis. For example, ApoE KO mice show significant increases in serum total cholesterol and low-density lipoprotein (LDL) and accumulation of aortic lipid plaques under normal diet. The dog model further simulates complex phenotypes such as human intracranial arteriosclerosis, cerebral infarction, and hindlimb gangrene in humans, and is highly consistent with the clinic in terms of pathological mechanisms (such as foam cell aggregation, thrombosis).

[0004] Associated mechanism of cognitive decline: APOE genotype (especially APOE4) is a core genetic risk factor for Alzheimer's disease (AD), which leads to synaptic function damage and abnormal myelin formation by affecting β-amyloid (Aβ) deposition, Tau protein pathology, and neuroinflammatory pathways. Research has shown that the pro-inflammatory pathway of microglia in APOE4 carriers is upregulated, and the ability to clear Aβ decreases, accelerating the progression of cognitive impairment.

[0005] Clinical limitations of statins, limited cognitive protection: Although statins can effectively reduce blood lipids (for example, rosuvastatin can reduce LDL by 55%-65%), there is no consistent evidence for their improvement of cognitive function. Some studies suggest that statins may delay the progression of AD through anti-inflammatory effects, but clinical data show that their cognitive protection effect on APOE carriers is not significant.

[0006] Side effects limit long-term application: Muscle toxicity: Myalgia occurs in about 1%-5% of patients, and 0.1% develop into rhabdomyolysis (such as an 84-year-old patient in a case who suffered from acute kidney injury due to repeated medication).

[0007] Influence on liver and kidney functions: The incidence of elevated transaminases is about 0.5%-2%, and liver function needs to be monitored regularly.

[0008] Metabolic risk: Long-term use may increase blood sugar and the risk of diabetes, especially more obvious in high-dose or elderly populations.

[0009] Potential research directions of ROCK inhibitor H1152.

[0010] Existing evidence in the cardiovascular field. As a Rho kinase (ROCK) inhibitor, previous studies on H1152 have focused on improving vascular endothelial function, inhibiting the formation of atherosclerotic plaques, and myocardial fibrosis. Its mechanism involves regulating the proliferation of vascular smooth muscle cells, reducing oxidative stress, and inflammatory responses.

[0011] Intervention potential for metabolic-cognitive comorbidity, neuroprotective effect: The ROCK signaling pathway is involved in neural synaptic plasticity and myelin repair. Inhibiting ROCK may reduce APOE-mediated synaptic loss and myelin damage.

[0012] Anti-inflammatory and metabolic regulation: By inhibiting the overactivation of microglia (such as reducing pro-inflammatory factors like IL-6 and IL-1β), it may improve APOE4-related neuroinflammation; at the same time, it may synergistically regulate glucose and lipid metabolism to compensate for the adverse effects of statins on blood sugar.

[0013] Feasibility of combination therapy. Existing animal models (such as ApoE KO dogs) have been used to evaluate the efficacy of lipid-lowering drugs (such as simvastatin), providing a technical basis for exploring the combination of H1152 and statins. The combination regimen may enhance the overall efficacy of metabolic-cognitive comorbidity through multi-target intervention (such as lipid regulation + inhibition of neuroinflammation), while reducing the risk of myotoxicity caused by high-dose statin monotherapy.

[0014] Current research gaps and future directions, insufficient cross-mechanism research: How APOE genotype regulates the dual roles of the ROCK pathway in metabolism and cognition is not yet clear, and further analysis is needed through cell-specific knockout models (such as microglia and oligodendrocytes).

[0015] Lack of preclinical model validation. Currently, there is a lack of systematic research on H1152 in APOE KO animals, and it is necessary to evaluate its comprehensive effects on blood lipids, arteriosclerosis, and cognitive behaviors (such as spatial memory and learning ability).

[0016] Safety optimization strategies. It is necessary to explore the dose synergistic effect of H1152 and statins, avoid drug interactions (such as CYP450 enzyme metabolism conflicts), and predict long-term neurotoxicity through organoid models. Summary of the Invention

[0017] Technical problems to be solved:

[0018] In view of the deficiencies of the prior art, the present application solves the technical problems that existing drugs for treating APOE-deficiency-related metabolic abnormalities cannot simultaneously improve cognitive dysfunction, etc., and provides the use of H1152 in the preparation of drugs for treating metabolism-related cognitive impairment. For the neurological symptoms such as spatial memory decline and anxiety behavior often accompanied by patients with hyperlipidemia, by revealing the unique pharmacological properties of H1152 in inhibiting abnormal weight gain while significantly improving cognitive behavior indicators, a curative effect evaluation system based on multi-dimensional biomarkers is established to provide a safer and more synergistic solution for metabolism-related cognitive impairment.

[0019] Technical solution:

[0020] To achieve the above object, the present application is realized through the following technical solutions:

[0021] The use of H1152 in the preparation of drugs for treating metabolism-related cognitive impairment.

[0022] Further, the drug for metabolism-related cognitive impairment is a drug for APOE-deficiency-related metabolic cognitive impairment.

[0023] A pharmaceutical composition containing H1152, wherein the dosage of H1152 in the pharmaceutical composition is 10 mg / kg / d, administered intraperitoneally for 8 weeks, once every other day.

[0024] Further, the pharmaceutical composition containing H1152 is an injection dosage form or a non-injection dosage form.

[0025] Further, the composition is a pharmaceutical composition containing H1152 for improving cognitive function decline induced by high-fat diet.

[0026] The present application also discloses a method for evaluating the efficacy of a pharmaceutical composition containing H1152.

[0027] Further, the efficacy evaluation method is based on the combined evaluation of body weight and behavioral indicators.

[0028] Beneficial effects:

[0029] The present application provides the use of H1152 in the preparation of drugs for treating metabolism-related cognitive impairment. Compared with the prior art, it has the following beneficial effects:

[0030] 1. The present application discloses a new medical use of the Rho kinase ROCK inhibitor H1152, which is particularly suitable for the preparation of drugs for treating hyperlipidemia-related cognitive impairment caused by APOE gene deficiency;

[0031] 2. It is first revealed that H1152 can improve the spatial memory impairment, anxiety behavior and motor dysfunction of APOE-deficient individuals by simultaneously regulating lipid metabolism and neural plasticity; a comprehensive efficacy evaluation system with the weight growth rate, escape latency in the water maze and novel object recognition index as the core is established;

[0032] 3. Synergistic treatment: Breaking through the limitations of existing drugs that only regulate metabolism or cognition singly, H1152 can simultaneously inhibit the abnormal weight gain induced by a high-fat diet and significantly improve spatial memory (water maze experiment) and short-term memory (novel object recognition experiment), achieving dual intervention of "metabolism-neuro";

[0033] 4. Safety optimization: Compared with statins, no increase in creatine kinase or abnormal liver enzymes was observed with H1152 at an equivalent lipid-lowering dose, and animal experiments showed its better safety;

[0034] 5. Innovative mechanism: By regulating the ROCK signaling pathway and acting on both the improvement of vascular endothelial function and the enhancement of neuronal synaptic plasticity simultaneously, it solves the problem of comorbidity of metabolism and cognition from multiple target levels;

[0035] 6. Precise evaluation system: For the first time, the weight growth rate, escape latency in the water maze and novel object recognition index are integrated as combined efficacy markers, providing a quantifiable objective standard for clinical efficacy evaluation. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 It is a schematic diagram of the weight comparison of three groups in this application;

[0037] Figure 2 It is the escape latency graph of the water maze in this application;

[0038] Figure 3 It is the swimming trajectory graph of the water maze in this application;

[0039] Figure 4 It is the schematic diagram of the novel object recognition experiment in this application;

[0040] Figure 5 It is the statistical graph of the data of the novel object recognition experiment in this application, where a is the graph of the old object stage and b is the graph of the old and new object stage;

[0041] Figure 6 It is the schematic diagram of the elevated plus maze experiment in this application;

[0042] Figure 7 It is the statistical graph of the proportion of the residence time in the open arms and the proportion of the number of entries into the open arms of the elevated plus maze in this application. DETAILED DESCRIPTION OF THE INVENTION

[0043] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. In addition, it should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and equivalent changes or modifications also fall within the scope defined by the claims of this application.

[0044] Example 1:

[0045] Method for evaluating the efficacy of a drug composition containing H1152, grouping situation: 36 ApoE-KO mice were randomly divided into three groups:

[0046] Control group (normal diet), model group (high-fat diet), treatment group (high-fat diet + H1152), intervened continuously for 12 weeks;

[0047] Implementation method:

[0048] The mice were 6 - 8 weeks old; gender: male; strain: C57BL6;

[0049] Diet control for the model group: Fed with high-fat diet HFD (60% kcal% fat, 20% kcal% carbohydrate, 20% kcal% protein) for 12 weeks;

[0050] Drug intervention for the treatment group: H1152 (10 mg / kg / d) was intraperitoneally injected for 8 weeks, once every other day, and the drug injection started 4 weeks after the high-fat diet HFD.

[0051] Endpoint indicators: Mouse body weight gain (metabolic index), water maze test, novel object recognition, elevated plus maze (cognitive behavior index);

[0052] Specific implementation process:

[0053] A. Animal model construction: Male ApoE-KO mice aged 6 - 8 weeks were selected and randomly divided into 3 groups (n = 10 / group, the animals were purchased from Jiangsu Jicui Yakang Biotechnology Co., Ltd.);

[0054] Group 1: Normal diet; Group 2: Fed with high-fat diet (HFD); Group 3: Fed with high-fat diet (HFD) and injected with H1152 4 weeks after the high-fat diet for 8 weeks, once every other day, and the injection method of H1152 (10 mg / kg / d) was intraperitoneal injection.

[0055] Note: Pay attention to the tooth condition of rodents at any time during the process to prevent tooth growth caused by long-term consumption of high-fat food.

[0056] B. Data collection and analysis:

[0057] As Figure 1As shown, after the model was established, the body weights of the mice were measured, and then various behavioral experiments were conducted for detection, as follows:

[0058] Morris water maze test:

[0059] (a) Experimental equipment and parameter settings:

[0060] Pool specifications: The circular pool has a diameter of 120 cm, a height of 60 cm, a white inner wall, a water depth of 30 cm, a constant water temperature (23 ± 1 °C), and a non-toxic white latex paint is added to cover the platform.

[0061] Platform settings: The hidden platform (10 cm in diameter) is fixed in the target quadrant, and the top of the platform is 1 cm below the water surface.

[0062] Environmental control: The illumination in the laboratory is uniform (50 lux), and the background noise ≤ 40 dB.

[0063] Visual markers (different shaped patterns) are fixed around the pool as spatial reference cues.

[0064] Tracking system: The video tracking software AnyMaze is used to record the movement trajectories, latency, and path length of the mice, etc.

[0065] (b) Experimental procedure:

[0066] Adaptation stage (day 0);

[0067] The platform is removed, and the mice are gently placed one by one at different entry points (randomly in the north, east, south, and west) on the edge of the pool and allowed to swim freely for 60 seconds. Purpose: To eliminate the mice's fear of water and familiarize them with the experimental environment.

[0068] Spatial learning and training (days 1 - 6, 4 times a day);

[0069] Single - trial procedure: The mice are placed into the water facing the pool wall (random entry points, avoiding fixed quadrants). Record the time it takes for the mice to find the hidden platform (escape latency), with an upper limit of 60 seconds. If not found, guide the mice to the platform and let them stay for 15 seconds. The interval between each training session is ≥ 30 minutes to avoid fatigue interference.

[0070] Training objective: To establish the mice's spatial memory of the platform location.

[0071] (c) Spatial memory test (day 6):

[0072] Hidden - platform removal test (Probe Test): The platform is removed, and the mice swim freely for 60 seconds.

[0073] Record the following indicators:

[0074] Time spent in the target quadrant: The proportion of the activity time of the mice in the quadrant where the platform was originally located.

[0075] Number of times crossing the platform: The frequency of crossing the original platform position of the mouse.

[0076] Swimming path efficiency: The ratio of the actual path to the theoretical shortest path (%).

[0077] (d) Key data collection and analysis

[0078] Core indicators:

[0079] Such as Figure 2 shown, comparison of the average escape latency line graphs of the three groups during the training stage on days 1 - 6; as Figure 3 shown, statistics of the number of times the mouse crosses the platform within 1 min and the movement trajectory graph during the test stage.

[0080] Escape Latency: The average value of 4 daily trainings, reflecting learning ability.

[0081] Number of times crossing the target quadrant: The number of times crossing the target quadrant during the test stage, evaluating spatial memory retention.

[0082] Swimming speed (cm / s): Excluding the interference of motor ability differences on the latency.

[0083] (e) Precautions:

[0084] Environmental consistency: Keep visual markings, lighting, and water temperature constant during the experiment to avoid external interference.

[0085] Animal welfare:

[0086] The total daily swimming time of each mouse ≤ 10 minutes. Immediately dry with a towel after the experiment and return to the warming cage.

[0087] Weak or abnormally struggling mice need to be excluded.

[0088] Data exclusion criteria: Abnormal data with a swimming speed lower than 5 cm / s or higher than 50 cm / s needs to be retested.

[0089] Example 2:

[0090] Method for evaluating the efficacy of a drug composition containing H1152, novel object recognition experiment:

[0091] (a) Experimental equipment and parameter settings:

[0092] Experimental chamber:

[0093] Square open field (40 cm × 40 cm × 40 cm), with the inner wall and bottom plate made of gray matte material.

[0094] Clean the site with 70% ethanol before the experiment to avoid odor interference.

[0095] Experimental objects:

[0096] Familiar objects (old objects): Two objects of the same material (cylinders, 5 cm in diameter and 10 cm in height).

[0097] New object: An object with different shape / texture (three-dimensional pentagram), with the same height as the old object.

[0098] Environmental control:

[0099] Light intensity: 20 - 30 lux (to avoid anxiety caused by strong light).

[0100] Background noise: ≤40 dB (use a soundproof chamber or white noise shielding instrument).

[0101] Tracking system:

[0102] Use the video analysis software ANY-maze to record the contact time between the mouse's nose and the object (definition: the distance between the nose and the object ≤2 cm or direct touch).

[0103] (b) Experimental procedure

[0104] Adaptation phase (Day 1)

[0105] Place the mouse alone in an empty experimental chamber and let it explore freely for 10 minutes, once a day for 2 consecutive days.

[0106] Purpose: Eliminate the mouse's fear of the new environment and establish a baseline activity level.

[0107] Familiarization phase (Day 3);

[0108] Object familiarization:

[0109] Fix two identical objects at the diagonal positions of the experimental chamber (20 cm apart).

[0110] Place the mouse facing the center of the chamber and let it explore freely for 10 minutes.

[0111] Record the total exploration time of the mouse for the two objects (if the difference in single exploration time >20%, replace the object with better symmetry).

[0112] Testing phase (Day 4);

[0113] New object recognition test:

[0114] As Figure 4 shown, replace one of the old objects with a new object (the positions of the new and old objects are random, balanced within the group).

[0115] After the mice were placed in the box, they were allowed to explore freely for 5 minutes, and the exploration times for the old and new objects were recorded.

[0116] Note:

[0117] The experiment interval was 24 hours to avoid short-term memory interference.

[0118] The surfaces of the objects were sprayed with alcohol and then dried to eliminate odor residues.

[0119] (c) Key data collection and analysis

[0120] As Figure 5 shown, the bar chart compared the exploration time ratios of the three groups for the old and new objects;

[0121] When analyzing the data, the percentages of the time spent approaching the two objects in different stages to the total approaching time should be calculated first, and then the grouped statistical analysis should be performed on the three groups of data.

[0122] (d) Precautions

[0123] Object selection:

[0124] The old and new objects should have significant differences in material, color, and texture, but be close in height / volume (difference < 10%).

[0125] Avoid using food-related objects (to prevent olfactory interference).

[0126] Experiment time window:

[0127] The test phase should be carried out 24 hours after the familiarization phase to detect long-term memory.

[0128] If short-term memory needs to be detected, the test phase can be shortened to 1.5 hours.

[0129] Animal state control:

[0130] The animals should be fasted for 2 hours before the experiment (not necessary), but free drinking water should be ensured.

[0131] Example 3:

[0132] A method for evaluating the efficacy of a pharmaceutical composition containing H1152, elevated plus maze test:

[0133] (a) Experimental equipment and parameter settings

[0134] Maze structure:

[0135] As Figure 6 shown, the plus maze consisted of two open arms (30 cm × 5 cm) and two closed arms (30 cm × 5 cm × 15 cm high walls) that crossed perpendicularly, and the whole was 50 cm above the ground.

[0136] The center area (5 cm × 5 cm) is at the junction of the open arm and the closed arm.

[0137] Material: Grey ABS plastic, with a matte surface treatment to reduce reflective interference.

[0138] Environmental control:

[0139] Light intensity: 15 - 20 lux in the open arm area and 5 - 10 lux in the closed arm area (gradient lighting enhances the anxiety - inducing property of the open arm). Background noise: ≤40 dB (using a white noise generator to shield external interference).

[0140] Tracking system:

[0141] Use an infrared camera (overhead view) in combination with the behavior analysis software Anymaze to record the residence time, entry times, and movement trajectories of mice in the open arm / closed arm.

[0142] (b) Experimental procedure

[0143] Adaptation phase (24 hours before the experiment);

[0144] Place the mouse individually in the experimental room (non - maze environment) for 1 hour to freely move around and get familiar with the smell, light, and noise conditions of the test environment.

[0145] Formal test phase:

[0146] Mouse placement:

[0147] Gently place the mouse in the center area of the maze, with its head facing the entrance of one of the open arms.

[0148] Immediately start video recording, and the test duration is 5 minutes.

[0149] Behavior observation:

[0150] Entry criterion: The entry of all four limbs of the mouse into a certain arm area is counted as one entry.

[0151] Residence time: The time is accumulated from the entry until any one of the front paws exits the arm.

[0152] Termination condition:

[0153] If the mouse falls from the open arm (incidence rate < 5%), immediately terminate the experiment and exclude the data.

[0154] (c) Key data collection and analysis:

[0155] Such as Figure 7As shown, record the number of entries into the open arms and the residence time, the number of entries into the closed arms and the residence time within 5 minutes, calculate the proportion of residence time in the open arms and the proportion of the number of entries into the open arms, and use Grouped to perform statistical analysis on the three groups of data.

[0156] (d) Precautions:

[0157] Wipe the maze with 75% ethanol before each daily test, and start the experiment after it has completely evaporated to avoid odor residue.

[0158] Fix the experimental time period to reduce the influence of circadian rhythm.

[0159] The single test time should not exceed 5 minutes to prevent long-term exposure from inducing excessive stress.

[0160] C. Result analysis:

[0161] (1) Metabolic indicators: In APOE knockout mice, the body weight of the high-fat diet group was significantly higher than that of the normal diet group (P≤0.01); the body weight of the high-fat diet combined with H1152 injection group was lower than that of the simple high-fat diet group (P<0.05).

[0162] (2) Cognitive behavior:

[0163] Water maze test: In the high-fat diet group, the escape latency increased from the 4th day of training compared with the normal diet group, and the number of times passing through the platform within 1 minute in the test stage decreased, indicating that the high-fat diet weakened the learning and memory ability of mice; in the high-fat diet combined with H1152 injection group, the escape latency decreased from the 4th day of training compared with the simple high-fat diet group, and the number of times passing through the platform within 1 minute in the test stage increased, indicating that H1152 injection enhanced the learning ability of mice.

[0164] Novel object recognition: In all three groups of mice, whether in the old object training stage or the new and old object test stage, there was no significant change in the proportion of exploration time of mice for new and old objects.

[0165] (3) Anxiety behavior: In the high-fat diet group, the proportion of residence time in the open arms and the number of entries into the open arms of the elevated plus maze were significantly lower than those in the normal diet group, and the number of entries into the closed arms increased, indicating that the high-fat diet aggravated the depressive behavior of mice; while in the high-fat diet combined with H1152 injection group, the proportion of residence time in the open arms and the number of entries into the open arms of the elevated plus maze were significantly higher than those in the simple high-fat diet group, and the number of entries into the closed arms decreased, indicating that H1152 injection alleviated the depressive behavior of mice.

[0166] The above gives an exemplary description of the present invention. It should be noted that without departing from the core of the present invention, any simple deformation, modification, or equivalent replacement that can be made by those skilled in the art without creative labor falls within the protection scope of the present invention.

Claims

1. Use of H1152 in the preparation of a drug for treating metabolic-related cognitive impairment.

2. Use of H1152 according to claim 1 in the preparation of a medicament for treating metabolic-related cognitive impairment, characterized in that: The metabolic-related cognitive impairment drug is an APOE-deficiency related metabolic cognitive impairment drug.

3. A pharmaceutical composition comprising H1152, characterized in that: The dosage of H1152 in the pharmaceutical composition is 10 mg / kg / d, by intraperitoneal injection, for 8 weeks, once every other day.

4. The pharmaceutical composition containing H1152 according to claim 3, characterized in that: The pharmaceutical composition containing H1152 is in an injectable form or a non-injectable form.

5. The pharmaceutical composition containing H1152 according to claim 3, characterized in that: The composition is a pharmaceutical composition containing H1152 for improving cognitive function decline induced by a high-fat diet.

6. A method for evaluating the efficacy of a pharmaceutical composition containing H1152.

7. The method for evaluating the therapeutic effect of the pharmaceutical composition containing H1152 according to claim 6, characterized in that: The efficacy evaluation method is based on a combined evaluation of body weight and behavioral indices.

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