Medicine for treating learning and memory impairment of Alzheimer disease and application

Abstract

The invention provides a medicine for treating learning and memory impairment of Alzheimer's disease and application. The medicine at least comprises PF-4708671, and the medicine at least comprises The learning and memory ability is tested and evaluated by using two AD mice as models, a 5xFAD transgenic mouse can simulate the advanced neuropathological process and behavioral performance of an AD patient in the early stage, and an A [beta] O injection mouse model not only can simulate abnormal behaviors of AD diseases, but also can simulate the advanced neuropathological process and behavioral performance of the AD patient in the later stage. A beta O is injected into a mouse model, and an injection brain region is selected to control ABO to intervene in a specific brain region, so that PF-4708671 can penetrate through a blood brain barrier to reach a target disease site, a new treatment direction is provided for AD (Alzheimer's disease), and particularly, a new drug selection is provided for learning and memory disorder caused by the Alzheimer's disease.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a medicine for treating learning and memory disorders caused by Alzheimer's disease and its application. Background Art

[0002] Alzheimer's disease (AD) is the most common neurodegenerative disease in the world. It is a degenerative lesion of the central nervous system characterized by progressive cognitive dysfunction and behavioral impairment. Its typical pathological features are neuritic plaques formed by the deposition of beta amyloid (Aβ) and neurofibrillary tangles (NFTs) formed by the phosphorylation of Tau protein, which eventually lead to functional damage of synapses and loss of neurons, further manifested as learning and memory impairment. However, the molecular signal transduction mechanism of AD pathology is still unclear, which has hindered the effective treatment of the disease. In particular, the therapeutic drugs in the prior art are only effective for some symptoms of AD, and the effect of use is not ideal.

[0003] The existing technology usually designs and develops corresponding drugs for treating learning and memory disorders in Alzheimer's disease based on different pathological hypotheses, mainly including: ① Cholinesterase inhibitors: tacrine, donepezil, rivastigmine, galantamine; ② NMDA receptor antagonists: memantine; ③ Treatment for Aβ: β-secretase inhibitors, γ-secretase inhibitors, aducanumab; ④ Treatment for Tau: Semorinemab, gosuranemab, zagotenemab; ⑤ Anti-infection treatment: non-steroidal anti-inflammatory drugs; ⑥ Treatment targeting the brain-gut axis: Ganlutena capsule. However, the above drugs basically have a certain degree of side effects and adverse reactions, and some drugs even show no obvious efficacy in clinical trials; further, since the above drugs do not have the effect of curing the disease, it is urgent to develop an effective and safe drug for the treatment of learning and memory disorders in Alzheimer's disease, which is of great significance.

[0004] On the other hand, PF-4708671 (2-[[4-(5-ethyl-4-pyrimidinyl)-1-piperazinyl]methyl]-6-(trifluoromethyl)-1H-benzimidazole) is a piperazinyl pyrimidine analog that is a specific inhibitor of S6K1, but PF-4708671 does not significantly inhibit the activity of the closely related S6K2 isoform, or other AGC kinases. PF-4708671 inhibits S6K1-mediated phosphorylation of S6 proteins in response to IGF-1 (insulin-like growth factor 1), and has no effect on PMA-induced phosphorylation of substrates that are highly related to RSK (p90 ribosomal S6 kinase) and MSK (mitogen and stress activated protein kinase) kinases. PF-4708671 induces phosphorylation of the T-loop and hydrophobic motifs of S6K1, an effect that is dependent on mTORC1 (mTOR complex 1). In the prior art, PF-4708671 has anti-tumor activity and can inhibit tumor cell proliferation in vitro and in vivo, such as Chinese invention patent CN110538182A; in addition, PF-4708671 can also be used in strategies to treat insulin resistance, and PF-4708671 has been shown to improve synaptic structure and functional defects in epilepsy.

[0005] The prior art also discloses that PF-4708671 can improve the memory level of Parkinson's disease mice in new object recognition tests. However, the above research mechanism is not the same as the pathology of Alzheimer's disease. Parkinson's disease and Alzheimer's disease belong to two different neurological diseases. Parkinson's disease is a chronic progressive neurodegenerative disease that mainly affects the dopaminergic neurons in the substantia nigra of the midbrain, resulting in a decrease in dopamine levels, which in turn causes movement disorders; while Alzheimer's disease is a chronic neurodegenerative disease that mainly affects the cerebral cortex, especially the hippocampus of the temporal lobe and parietal lobe, resulting in impaired cognitive functions such as memory, thinking, and behavior. Based on different pathological processes, the drugs used for treatment are completely different. Parkinson's treatment drugs are mainly levodopa, dopamine receptor agonists, etc., while Alzheimer's disease drugs are mainly cholinesterase inhibitors, NMDA receptor antagonists, etc. The existing research on PF-4708671 does not involve the study of the pathological process of Alzheimer's disease, nor has PF-4708671 been used in the treatment of Alzheimer's disease, especially no relevant reports on the treatment of learning and memory disorders caused by AD. Summary of the invention

[0006] In view of the technical problems existing in the prior art, the present invention discloses a medicine for treating learning and memory disorders caused by Alzheimer's disease and its application.

[0007] To achieve the above object, the present invention provides a use of PF-4708671 in a drug for treating learning and memory disorders in Alzheimer's disease.

[0008] PF-4708671 (2-[[4-(5-ethyl-4-pyrimidinyl)-1-piperazinyl]methyl]-6-(trifluoromethyl)-1H-benzimidazole), the structural formula is:

[0009]

[0010] Preferably, PF-4708671 can penetrate the blood-brain barrier and enter the DG region of the hippocampus to reduce the phosphorylation level of RNF168.

[0011] As one of the purposes of the invention, the present invention also provides a drug for treating learning and memory disorders in Alzheimer's disease, comprising at least PF-4708671.

[0012] Preferably, PF-4708671 can be used as the sole active ingredient in a drug for treating learning and memory disorders in Alzheimer's disease.

[0013] Studies have shown that based on the pharmacokinetics and blood-brain distribution spectrum of PF-4708671, PF-4708671 can be detected in the brain after intraperitoneal injection of 32 mg / kg PF-4708671 in mice. PF-4708671 is able to pass through the blood-brain barrier, ensuring that the drug penetrates into the brain parenchyma to reach the target disease site, thereby enabling it to be used in the treatment of neurological diseases.

[0014] Preferably, the drug may further include pharmaceutically acceptable auxiliary ingredients.

[0015] Furthermore, PF-4708671 can also be used as an active ingredient in drugs for treating complications of Alzheimer's disease.

[0016] Preferably, the Alzheimer's disease complications include Alzheimer's disease combined with one or more of tumors, diabetes, epilepsy, and Parkinson's disease.

[0017] Based on the above technical scheme, the specific scheme adopted by the present invention includes: selecting 4-5 month old male 5xFAD mice and control wild-type mice, intraperitoneally injecting PF-4708671 or the corresponding blank control (vehicle) for 1 hour, and evaluating the learning and memory ability through Y maze test, new object recognition test, new position recognition test and Morris water maze test, and detecting the motor ability by rotating rod fatigue test; further, selecting 6-8 week old male Aβ oligomer (AβO) injected mice and wild-type mice injected with the corresponding vehicle, repeating the above steps for detection; the results showed that PF-4708671 can improve the learning and memory ability of 5xFAD mice and AβO injected mice.

[0018] The technical effects of the technical solution of the present invention are as follows:

[0019] (1) The present invention uses two AD mouse models to test and evaluate learning and memory abilities. The 5xFAD transgenic mouse can simulate the neuropathological process and behavioral manifestations of late-stage AD patients at an early stage, providing a suitable disease environment for the development of AD therapeutic drugs. The AβO-injected mouse model can not only simulate the abnormal behavior of AD symptoms and has the characteristics of strong operability, but PF-4708671 can also improve the learning and memory abilities of 5xFAD mice and AβO-injected mice.

[0020] (2) The present invention discloses for the first time the use of PF-4708671 in the treatment of AD. PF-4708671 can penetrate the blood-brain barrier to ensure that the drug penetrates into the brain parenchyma to reach the target disease site. It is suitable for neurological diseases and provides a new treatment direction for AD, especially a new medication option for learning and memory disorders caused by Alzheimer's disease.

[0021] (3) Through the technical solution of the present invention, it is revealed that PF-4708671 can improve the learning and memory disorders caused by Alzheimer's disease, and further has potential therapeutic effects in diseases such as tumors, diabetes, epilepsy, Parkinson's disease, etc., and has the advantage of wide efficacy, thereby providing a new medication regimen for the treatment of patients with AD combined with the above diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1A-1E This is a graph showing the test results of using PF-4708671 to improve the learning and memory abilities of 5xFAD mice in a typical embodiment of the present invention.

[0023] Figure 2A-2E This is a graph showing the test results of PF-4708671 improving the learning and memory abilities of AβO-injected mice in a typical embodiment of the present invention.

[0024] Figure 3A and Figure 3B Representative immunoblot images and statistical analysis graphs of pS60-RNF168 and total RNF168 levels in the hippocampal DG of 5xFAD mice after injection of PF-4708671 in a typical embodiment of the present invention are shown, respectively.

[0025] Figure 3C and Figure 3D They are representative immunoblot images and statistical analysis graphs of pS60-RNF168 and total RNF168 levels in the hippocampal DG of AβO model mice after injection of PF-4708671 in a typical embodiment of the present invention. DETAILED DESCRIPTION

[0026] To make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the protection scope of the present invention.

[0027] The present invention uses two AD mouse models, in which 5xFAD mice develop Aβ deposition plaques at 2 months of age. Learning and memory-related behavioral disorders begin to appear at 4-5 months of age. 5xFAD transgenic mice can simulate the neuropathological process and behavioral manifestations of late-stage AD patients at an early stage, providing a suitable disease environment for the development of AD therapeutic drugs. The AβO injection mouse model can not only simulate AD behavioral abnormalities, but also has stronger operability, which is reflected in the present invention as being able to intervene in specific brain areas.

[0028] The technical scheme of the present invention is further described in detail below through specific examples. The experimental methods in the following examples without specifying specific conditions are usually carried out under conventional conditions or under conditions recommended by the manufacturer.

[0029] Example 1

[0030] 1. Materials and Methods

[0031] 1.1 Experimental animals

[0032] Two AD mouse models were selected: 5xFAD and AβO-injected mice. 5xFAD mice have a C57BL / 6J genetic background and can overexpress human amyloid precursor protein (APP) and presenilin protein-1 (PSEN1), which contains a total of 5 AD-related mutations: Swedish (K670N / M671L), Florida (1716V) and London (V717I) mutations in APP, and M146L and L286V mutations in PSEN1. 5xFAD mice were selected at 4-5 months of age, and C57BL / 6J mice of the same age were used as the control group. The AβO-injected mouse model used 6-8 weeks of age C57BL / 6J mice (purchased from Nanjing Jicui Pharmaceutical Kang Biotechnology Co., Ltd.), and C57BL / 6J mice of the same age injected with a blank control vehicle were used as the control group.

[0033] 1.2 AβO preparation and stereotaxic brain injection

[0034] Dissolve 1.0 mg of human Aβ1-42 (Shanghai Jie Nitai Biotechnology Co., Ltd., HA-42-T-2) lyophilized powder in 400 μL HFIP (Sigma, 920-66-1) and incubate at room temperature for 30 minutes until clear. The solution was divided into 4 EP tubes and dried peptide particles were obtained by vacuum centrifuge. After resuspending to 5 mM with DMSO (Solarbio, D8370), it was diluted to 0.1 mM with artificial cerebrospinal fluid without glucose. After incubation at 4°C for 24 hours, centrifugation (14000g, 10min, 4°C) was performed to obtain the supernatant, and the concentration was determined using the BCA method (Biyuntian, P0012). C57BL / 6J mice were anesthetized with ketamine (100 mg / kg, ip) combined with xylazine (10 mg / kg, ip) and fixed in a brain stereotaxic instrument. AβO (0.1 mM, 2.5 μL / side) or a blank control vehicle (a mixture of DMSO and artificial cerebrospinal fluid without glucose in the same proportion as the experimental group) was injected into the DG region of the hippocampus on both sides (anterior / posterior: -2.1 mm, medial / lateral: ±1.5 mm, dorsal / ventral: -2.1 mm) at a rate of 1 μμL / min using a microinjection pump. After the needle stayed in place for 2 minutes, the syringe was pulled out and the wound was sutured.

[0035] 1.3PF-4708671 (MedChemExpress, HY-15773)

[0036] When used, it was dissolved in physiological saline containing 17% DMSO and 10% Tween80 (mass: 50 mg / kg; volume: 5 ml / kg, i.β.) and injected 1 h before the experiment.

[0037] 1.4 Behavioral testing

[0038] The experimental groups were as follows: WT+veh, 5xFAD+veh and 5xFAD+PF groups were composed of C57BL / 6J or 5xFAD mice injected intraperitoneally with vehicle (saline containing 17% DMSO and 10% Tween80) or PF-4708671, respectively; veh+veh group was composed of C57BL / 6J mice injected stereotaxically with vehicle (see 1.2) and intraperitoneally with vehicle (saline containing 17% DMSO and 10% Tween80); AβO+veh group was composed of C57BL / 6J mice injected stereotaxically with AβO and intraperitoneally with vehicle (saline containing 17% DMSO and 10% Tween80); AβO+PF group was composed of C57BL / 6J mice injected stereotaxically with AβO and intraperitoneally with PF-4708671. All tests were conducted at the same time each day. Mice were placed in the test room 1 hour before the experiment to adapt to the environment. Different batches of mice were used for different behavioral tests. Each experimental group used at least 8 mice. Learning and memory-related behavioral tests were performed 1 hour after PF-4708671 drug injection treatment. The test method is as follows.

[0039] 1.4.1Y maze test (Ymaze)

[0040] The Y-shaped maze consists of three arms (30×6×15 cm), with an angle of 120 degrees between two adjacent arms and a movable partition in the middle. The mouse was placed at the end of any arm of the Y-shaped maze and allowed to explore freely for 10 min. The total number of times the mouse entered the maze arm and the number of alternating entries (the number of consecutive entries into the three arms of the Y-shaped maze) were recorded. The alternation rate (%) of each mouse was calculated as [(alternation number) / (total number of entries-2)]×100.

[0041] 1.4.2 Novel Object Recognition Test (NORT) and Novel Location Recognition Test (NPRT)

[0042] Stage 1: Place mice individually in an open behavior box (50×50×50cm) and allow them to explore freely for 10 minutes; Stage 2: Place two identical objects in the behavior box and allow the mice to explore again for 10 minutes; Stage 3: ① Replace one of the objects with a new object or move it to a diagonal position and allow the mice to explore again for 10 minutes. The exploration behavior record starts when the mouse approaches or touches the identified object with its nose and ends when it leaves. The behavior of the mouse gnawing the identified object is not considered as exploration behavior and is not recorded.

[0043] The Anymaze system was used to measure the time mice spent exploring the new object and the non-replaced object. The discrimination index was (new object or location exploration time - old object or location exploration time) / total exploration time.

[0044] 1.4.3 Rotarod fatigue test

[0045] During the training phase, the fatigue rotarod was set to continuously accelerate the shaft to 50 rpm / min within 309 s, and the mice were placed on the shaft. After 1 min of adaptation, the rotarod was started and the time when the mice fell was recorded. Training was performed 3 times a day, with an interval of 20 min each time. On the third day, the test was performed, and each mouse was tested twice, and the falling time was recorded and the average value was taken.

[0046] 1.4.4Morris water maze

[0047] On the first day, each mouse was tested four times in a water maze (circular pool; diameter: 150 cm; height: 50 cm, water depth: 30 cm). Each time, the mouse entered the water from a different quadrant (SW, NW, NE, SE). Titanium dioxide was sprinkled in the water to make the pool turbid, and the mice were trained to find a visible resting platform (1 cm above the water). For the next 5 consecutive days, the mice were randomly allowed to enter the water from 4 different quadrants until they found a hidden resting platform (1 cm under the water). The time from entering the water to climbing onto the platform was recorded, and the average of the 4 times was the latency. If the mouse did not find the platform within 60 seconds, the experiment was terminated, and it was guided to the platform and stayed for 20 seconds before being picked up. On the 6th day, the platform was removed and a fixed 60-second swimming time was given. The time spent in the quadrant where the original platform was located, the number of times it crossed the platform area, and the swimming speed were recorded.

[0048] 1.4.5 PF-4708671 inhibits S6K1 activity test

[0049] 4-5 month old 5xFAD mice and control C57BL / 6J mice were selected. PF-4708671 (50 mg / kg; 5 mL / kg, ip) or vehicle (17% DMSO and 10% Tween80 in saline) was injected intraperitoneally 1 hour later. Then, ketamine (100 mg / kg, ip) combined with xylazine (10 mg / kg, ip) was used for anesthesia, and the DG region of the hippocampus was dissected under a microscope.

[0050] An appropriate amount of lysis buffer (containing 150 mM NaCl, 30 mM HEPES, 10 mM NaF, 1% v / v TritonX-100, 0.01% w / v SDS and protease inhibitors, pH 7.5) was added to the homogenate and ultrasonically lysed for 30 seconds. After centrifugation (16000g, 4°C, 10 min), the supernatant was collected and the protein concentration was determined by the BCA method.

[0051] Part of the supernatant was taken as whole cell lysate (WCL), mixed with 4× protein loading buffer, and incubated at 50°C for 15 min. Another appropriate amount of supernatant (containing 2 mg protein) was added with 1 μL RNF168 antibody (ABclonal Technology, A3556) and incubated at 4°C overnight. Then, protein A / G agarose beads (Changzhou Tiandirenhe Biotechnology, rProtein A / G MagPoly Beads, SM01505) were added and incubated at 4°C for 1 h. The agarose beads were then washed three times with lysis buffer, mixed with 1× protein loading buffer, incubated at 50°C for 15 min, separated from the WCL sample on a 10% SDS polyacrylamide gel, and then transferred to a nitrocellulose membrane. After soaking in blocking solution (5% skim milk powder dissolved in TBST buffer) for 1 hour, pRNF168 (1:1000, ABclonal Technology, AP1219), RNF168 (1:1000, ABclonal Technology, A3556) and GAPDH (1:10000, HuaAn Bio, ET1601-4) antibodies were diluted with 1% BSA-TBST buffer and incubated overnight at 4°C. After rinsing with TBST, rabbit secondary antibodies (1:10000, Wuhan Sanying Bio Proteintech, SA00001-2) diluted with 1% BSA-TBST buffer were incubated at room temperature for 1 hour; after rinsing with TBST buffer, developer (ultrasensitive ECL chemiluminescence kit, Beyotime, P0018M) was evenly added to the membrane surface, exposed using a gel imager, and grayscale value analysis of the target bands was performed using the Image J image processing system.

[0052] In addition, 6-8 week old AβO injected mice and C57BL / 6J mice injected with vehicle stereotaxic (see 1.2 for methods) were selected and the above steps were repeated for further testing. At least 3 mice were used in each experimental group.

[0053] Experimental results analysis

[0054] According to the technical solution of Example 1 above, the test results of PF-4708671 for improving the learning and memory ability of 5xFAD mice are shown in Figure 1A-1E .

[0055] Figure 1AThe results of the Y-maze test in the figure show that compared with the WT+veh group, the alternation rate of the 5xFAD+veh group decreased, while the alternation rate of the 5xFAD+PF group recovered. Among them, WT+veh, n=8; 5xFAD+veh, n=8; 5xFAD+PF, n=9. One-way ANOVA combined with post-hoc Tukey's comparisons test was used to compare the significance of the differences among the groups; among them, * represents P<0.05; ** represents P<0.01.

[0056] like Figure 1B and Figure 1C As shown in the novel position recognition test (NPRT) and the novel object recognition test (NORT), the discrimination index of the 5xFAD+veh group decreased compared with the WT+veh group in both NPRT and NORT, while the discrimination index of the 5xFAD+PF group improved compared with the 5xFAD+veh group. Among them, WT+veh, n=8; 5xFAD+veh, n=9; 5xFAD+PF, n=9. One-way ANOVA combined with Tukey's post hoc test was used to compare the significance of the differences among the groups. Among them, * represents P<0.05; *** represents P<0.001.

[0057] Figure 1D The results of the rotarod fatigue test shown in the figure show that the influence of the mouse movement difference on the experiment is excluded. Among them, WT+veh, n=8; 5xFAD+veh, n=9; 5xFAD+PF, n=9. One-way ANOVA was used to compare the significance of the differences among the groups. NS means no significant difference.

[0058] Figure 1EIn the Morris water maze test shown in the figure, the results showed that in the training phase, the latency of the 5xFAD+veh group to find the hidden platform was significantly prolonged compared with the WT+veh group, while the latency of the 5xFAD+PF group was significantly shortened compared with the 5xFAD+veh group; in the test phase, the time the 5xFAD+veh group stayed in the target quadrant and the number of times it crossed the platform area were significantly reduced compared with the WT+veh group, while the 5xFAD+PF group increased compared with the 5xFAD+veh group, and there was no significant difference in the swimming speed of the mice in each group. Among them, WT+veh, n=8; 5xFAD+veh, n=9; 5xFAD+PF, n=9, repeated two-way ANOVA was used to compare the significance of the difference in escape latency among the groups; KW test (Kruskal-Wallistest) combined with post-hoc Dunn's multiple comparisons test was used to compare the significance of the difference in the target quadrant residence time among the groups. One-way ANOVA combined with Tukey's post hoc test was used to compare the significance of the number of crossings. One-way ANOVA was used to compare the significance of the swimming speed among the groups. * represents P < 0.05; ** represents P < 0.01; NS represents no significant difference.

[0059] The above behavioral results indicate that PF-4708671 can improve the learning and memory abilities of 5xFAD mice.

[0060] Based on the test results of PF-4708671 improving the learning and memory ability of AβO injected mice in the above examples, see Figure 2A-2E .

[0061] See also Figure 2A Y-maze test. As shown in the figure, compared with the veh+veh group, the alternation rate of the AβO+veh group decreased, while the alternation rate of the AβO+PF group recovered. Among them, veh+veh, n=17; AβO+veh, n=14; AβO+PF, n=10. Statistical analysis methods are the same Figure 1A ; In the figures, *, P<0.05; **, P<0.01.

[0062] See also Figure 2B and Figure 2CIn the novel position recognition test (NPRT) and the novel object recognition test (NORT), as shown in the figure, in the NPRT and NORT, compared with the veh+veh group, the discrimination index of the AβO+veh group decreased, while the discrimination index of the AβO+PF group improved compared with the AβO+veh group; in the NRPT, veh+veh, n=13; AβO+veh, n=9; AβO+PF, n=11; in the NORT, veh+veh, n=14; AβO+veh, n=9; AβO+PF, n=10, the statistical analysis method was the same Figure 1B and Figure 1C ; Among them, * represents P < 0.05; *** represents P < 0.001.

[0063] See also Figure 2D The results of the rotarod fatigue test shown in the figure show that the influence of mouse movement differences on the experiment was eliminated. Among them, veh+veh, n=14; AβO+veh, n=11; AβO+PF, n=11; statistical analysis methods were the same as Figure 1D ,In the figure, NS means no significant difference.

[0064] See also Figure 2E , in the Morris water maze test shown in Figure , during the training phase, the latency of the AβO+veh group to find the hidden platform was significantly prolonged compared with the veh+veh group, while the latency of the AβO+PF group was significantly shortened compared with the AβO+veh group; during the test phase, the time the AβO+veh group stayed in the target quadrant and the number of times they crossed the platform area were significantly reduced compared with the veh+veh group, while the AβO+PF group increased compared with the AβO+veh group, and there was no significant difference in the swimming speed of the mice in each group; among them, veh+veh, n=25; AβO+veh, n=22; AβO+PF, n=10; the statistical analysis method was the same as Figure 1E Among them, the significance of the difference in the target quadrant residence time of each group was analyzed by one-way analysis of variance combined with Tukey's post hoc test, * represents P < 0.05; *** represents P < 0.001; NS represents no significant difference.

[0065] The above behavioral results indicate that PF-4708671 can also improve the learning and memory abilities of AβO-injected mice.

[0066] See also Figure 3A and Figure 3B , are representative immunoblot images and statistical analysis graphs of pS60-RNF168 and total RNF168 levels in the hippocampal DG of WT+veh, 5xFAD+veh, and 5xFAD+PF mice in this example, respectively. Figure 3C and Figure 3DRepresentative immunoblot images and statistical analysis of pS60-RNF168 and total RNF168 levels in the hippocampal DG of veh+veh, AβO+veh and AβO+PF mice in the examples are shown. As can be seen from the figure, PF-4708671 can reduce the phosphorylation level of RNF168 in the hippocampal DG of 5xFAD and AβO injected mice, while the total level of RNF168 does not change significantly. Therefore, it can be seen from the above results that RNF168 is a substrate of S6K1 (Xiaoduo Xie et al., Nat Cell Biol., 2018 March, 20(3): 320-331), and intraperitoneal injection of PF-4708671 can effectively inhibit the activity of S6K1 in the hippocampal DG of AD mice. WT+veh, n=4; 5xFAD+veh, n=4; 5xFAD+PF, n=4; veh+veh, n=3; AβO+veh, n=3; AβO+PF, n=3. One-way ANOVA combined with Tukey post hoc test was used to compare the significance of the differences among the groups. * represents P<0.05; ** represents P<0.01; *** represents P<0.001; NS represents no significant difference.

[0067] The above are only preferred embodiments of the present invention, which do not limit the protection scope of the present invention. For those skilled in the art, the present invention may have various modifications and changes. Within the spirit and principle of the present invention, changes, modifications, replacements, integrations and parameter changes to these embodiments by conventional substitutions or by being able to achieve the same functions without departing from the principles and spirit of the present invention all fall within the protection scope of the present invention.

Claims

1. Application of PF-4708671 in the treatment of learning and memory disorders in Alzheimer's disease. The structural formula of PF-4708671 is:

2. The use according to claim 1, characterized in that PF-4708671 can penetrate the blood-brain barrier and enter the DG region of the hippocampus, reducing the phosphorylation level of RNF168.

3. A drug for treating learning and memory disorders in Alzheimer's disease, characterized in that: Includes at least PF-4708671.

4. The drug according to claim 3, characterized in that PF-4708671 is the only active ingredient of the drug.

5. The drug according to claim 3, characterized in that: The medicament further comprises pharmaceutically acceptable auxiliary ingredients.

6. Application of PF-4708671 in the treatment of complications of Alzheimer's disease.

7. The use according to claim 6, wherein the complications of Alzheimer's disease include Alzheimer's disease combined with one or more of tumors, diabetes, epilepsy, and Parkinson's disease.

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