Application of SAA1 protein inhibitor in preparation of medicine for treating methylamphetamine addiction withdrawal syndrome

Abstract

The invention belongs to the cross technical field of biological medicine and neuropsychiatric medicine, and particularly relates to application of an SAA1 protein inhibitor in preparation of a medicine for treating methylamphetamine addiction withdrawal syndrome. The invention reveals that the mesenteric white adipose tissue plays a key role in anxiety, depression and other negative emotions induced by withdrawal of methylamphetamine by secreting SAA1 protein for the first time, and further obviously increases the drug foraging behavior and relapse risk. By inhibiting or reducing SAA1 protein expression or activity from mesenteric white fat, negative emotion related to withdrawal can be effectively relieved, so that psychological desire and relapse rate are reduced. The invention provides an inhibitor taking SAA1 protein as a target spot, an antibody, a gene silencing tool or application thereof. The inhibitor, the antibody and the gene silencing tool are used for preparing drugs or preparations for treating methylamphetamine addiction withdrawal syndrome and preventing relapse. The target and the treatment method provide a brand new way and strategy for material use disorder, especially clinical intervention of methamphetamine dependence.

Description

Technical Field

[0001] This invention belongs to the interdisciplinary field of biomedicine and neuropsychiatry, specifically relating to the application of SAA1 protein inhibitors in the preparation of drugs for treating methamphetamine addiction withdrawal syndrome. Background Technology

[0002] Methamphetamine (METH) addiction withdrawal syndrome refers to a series of clinical symptoms mainly characterized by mental and emotional symptoms that occur in long-term abusers after abruptly stopping or reducing the dosage. Its core manifestations are negative emotions (anhedonia, anxiety, depression), intense psychological cravings, and cognitive impairment (decreased attention, drowsiness, etc.), while physical symptoms are relatively mild. The course of the disease is usually divided into an acute withdrawal period (with the most severe emotional symptoms within 1 to 2 weeks after stopping the drug) and a protracted period (lasting for several weeks to several months, with residual anxiety, depression, and psychological cravings, which is a high-risk stage for relapse).

[0003] In the clinical treatment of methamphetamine addiction, relapse prevention is the biggest challenge. Studies show that even after compulsory or voluntary detoxification treatment, the annual relapse rate among methamphetamine addicts still exceeds 60%. The key factor contributing to this severe challenge stems primarily from the persistent and intense negative emotional state during long-term withdrawal, including anxiety, depression, anhedonia, and irritability (i.e., the aforementioned methamphetamine withdrawal syndrome). These negative emotions constitute the core driving force of "negative reinforcement"—individuals compulsively seek and relapse into the drug to alleviate or escape this unbearable negative emotional experience. Therefore, negative emotions during withdrawal are the core psychological motivation driving relapse behavior and a crucial link in the transformation of addiction from acute substance dependence to chronic relapsing encephalopathy.

[0004] Currently, because the withdrawal symptoms of METH withdrawal syndrome are centered on negative emotions (such as anxiety, depression, and anhedonia), doctors often try to treat the symptoms with conventional antidepressants (such as selective serotonin reuptake inhibitors, SSRIs) or anti-anxiety medications. However, these drugs are originally designed for primary mood disorders, and their pharmacological mechanisms of action mainly target the classic monoamine neurotransmitter system, making it difficult to cover the complex pathological processes involving peripheral-central interactions driven by METH neurotoxicity. Therefore, currently used drugs often have slow onset of action, low response rates, or even no effect in METH withdrawal patients. Summary of the Invention

[0005] To develop a novel treatment strategy for METH withdrawal syndrome and address the problems of slow onset of action, low response rate, or even ineffectiveness of existing clinically used drugs in METH withdrawal patients, this invention provides the application of an SAA1 protein inhibitor in the preparation of drugs for treating METH addiction withdrawal syndrome. To achieve the above objective, this invention employs the following technical solution.

[0006] This invention provides the application of SAA1 protein inhibitors in the preparation of drugs for treating METH addiction withdrawal syndrome. The SAA1 protein is serum amyloid A1.

[0007] SAA1 protein, as an acute-phase reactive protein, is abnormally highly expressed during METH withdrawal and enters the brain to activate microglia-mediated neuroinflammatory responses, affecting synaptic plasticity and ultimately exacerbating withdrawal symptoms such as anxiety, depression, and cravings. This invention, by specifically inhibiting the SAA1 protein, can block the neuroinflammatory cascade response at its source, rapidly alleviating core symptoms, thereby overcoming the shortcomings of traditional drugs that have low response rates or are ineffective due to dispersed targets, slow onset of action, or inability to block inflammatory pathways.

[0008] Furthermore, the drug exerts its effect by inhibiting or reducing the expression level or biological activity of SAA1 protein in mesenteric white adipose tissue.

[0009] Furthermore, the SAA1 protein is a human SAA1 protein or its mammalian homolog.

[0010] Furthermore, the mammal is selected from mice, rats, rabbits, dogs, pigs, or non-human primates.

[0011] Furthermore, the mammal is a mouse, and the amino acid sequence of the SAA1 protein is shown in SEQ ID NO.1, which is a precursor protein sequence containing a signal peptide: The sequence is: MKLLTGLVFCSLAVLGVSAQQWTAFISHEARQAGVQDMIRAYQDMKEANWKNSTVLVSKGNGDAAVLVPGGPEAWAAEVVSNAKENGIKAALTGRGEDSLADQAANKWGR SGKDPNRFRP KGLPDKY, where amino acids 1-18 (MKLLTGLVFC SLAVLGVSA) constitute the signal peptide sequence, and amino acids 19-122 (QQWTAFISHE... to the end) constitute the mature protein sequence. This sequence corresponds to UniProt accession number P05366.

[0012] Furthermore, the SAA1 protein inhibitor is at least one of the following: Antibodies or antigen-binding fragments that specifically bind to the SAA1 protein.

[0013] Small molecule inhibitors of the SAA1 protein.

[0014] mRNAs that target the SAA1 protein, including siRNA, shRNA, antisense oligonucleotides, or miRNA.

[0015] A CRISPR / Cas9 gene editing system targeting the gene encoding the SAA1 protein.

[0016] A carrier system for specifically delivering the SAA1 protein inhibitor to mesenteric white adipose tissue.

[0017] Furthermore, the vector system is an adeno-associated virus vector, which contains a white adipose tissue-specific promoter.

[0018] The white adipose tissue-specific promoter is either the mouse AP2 promoter or the human FABP4 promoter.

[0019] Furthermore, the adeno-associated virus vector is an AAV8 serotype vector (AAV-AP2 vector).

[0020] Furthermore, the shRNA of the mRNA targeting the SAA1 protein comprises a sense strand with a nucleotide sequence as shown in SEQ ID NO:2 and an antisense strand with a nucleotide sequence as shown in SEQ ID NO:3.

[0021] Furthermore, the antibody is a monoclonal antibody or a polyclonal antibody.

[0022] The monoclonal antibody is selected from humanized antibodies, fully human antibodies, and mouse monoclonal antibodies.

[0023] Furthermore, the drug is a liquid preparation.

[0024] Furthermore, the drug is used for local or systemic administration to the mesenteric white adipose tissue.

[0025] Furthermore, the METH addiction withdrawal syndrome includes anxiety, depression, irritability, or other negative emotional symptoms that occur during withdrawal.

[0026] Compared with the prior art, the present invention has the following beneficial effects: To develop a novel treatment strategy for METH withdrawal syndrome, this invention provides the application of a SAA1 protein inhibitor in the preparation of drugs for treating METH addiction withdrawal syndrome, thus offering a new treatment strategy for METH addiction withdrawal syndrome. The technical solution of this invention is based on the following original discovery: serum amyloid A1 (SAA1 protein) levels are significantly elevated in plasma during METH withdrawal, and its level is closely related to the intensity of withdrawal emotions. Further research revealed that peripheral mesenteric white adipose tissue is the main source of elevated SAA1 protein during withdrawal. Elevated SAA1 protein can enter the brain and act on SCARB1 receptors on neurons in the medial prefrontal cortex, thereby regulating synaptic plasticity and microglia activation, ultimately driving negative emotions and relapse behavior. By using an SAA1 protein inhibitor, this novel regulatory pathway of the "peripheral adipose-medial prefrontal cortex" neural axis can be blocked, fundamentally intervening in the generation of negative emotions and relapse tendency during withdrawal.

[0027] Based on the above mechanism, this invention can effectively block the neuropathological process mediated by the SAA1-SCARB1 signaling pathway by inhibiting the activity or expression of SAA1 protein, thereby solving the problem that existing drugs used in clinical practice often have slow onset, low response rate or even no effect in METH withdrawal populations. Attached Figure Description

[0028] Figure 1 This is a diagram showing the experimental results demonstrating that peripheral mesenteric white adipose tissue is the main source of SAA1 upregulation in the METH withdrawal model in this invention; wherein: Figure A shows the results of real-time quantitative PCR detection of SAA1 mRNA expression levels in peripheral tissues (including mesenteric white adipose tissue) and central brain regions; Figure B shows the results of enzyme-linked immunosorbent assay (ELISA) detection of SAA1 protein expression levels in peripheral tissues (including mesenteric white adipose tissue) and central brain regions; Image C is a representative image obtained by immunofluorescence staining to verify SAA1 expression in mesenteric white adipose tissue.

[0029] Figure 2 This is a diagram showing the experimental results of this invention demonstrating that peripheral intervention in the upregulation of SAA1 in mesenteric white adipose tissue can reverse METH withdrawal-related negative behaviors. Wherein:

[0030] A is the overall experimental flowchart, showing the timeline for adeno-associated virus injection containing SAA1-shRNA, establishment of the METH withdrawal model, and behavioral testing. B is a representative image of SAA1 knockdown in mesenteric white adipose tissue verified by immunofluorescence staining; Figure C shows the quantitative results of real-time quantitative PCR verification of the SAA1 knockdown effect in mesenteric white adipose tissue; D is the result of the open field experiment to detect the effect of the above AAV intervention on central region time (anxiety-like behavior); E is the result of the open field experiment to detect the effect of the above AAV intervention on total range of motion (motor ability); F is the result of the elevated cross maze experiment to detect the effect of the above AAV intervention on anxiety-like behaviors; G is the result of the tail suspension test to detect the effect of the above AAV intervention on depressive-like behavior; H represents the results of the forced swimming experiment used to detect the effects of the above-mentioned AAV intervention on depressive-like behaviors. Detailed Implementation

[0031] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments, but this should not be construed as limiting the invention. Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art, and the materials, reagents, etc. used in the following embodiments are commercially available unless otherwise specified.

[0032] The materials used in the following embodiments are as follows: (1) Reagents TRIzol lysis buffer, real-time quantitative PCR kit (SYBR Green), RIPA protein lysis buffer, BCA protein quantitative kit, rabbit-derived SAA1 polyclonal antibody, rabbit-derived GAPDH polyclonal antibody, HRP-labeled goat anti-rabbit secondary antibody, fluorescently labeled goat anti-rabbit secondary antibody, DAPI staining solution, sodium pentobarbital, methamphetamine (METH) were purchased from the National Narcotics Laboratory, physiological saline, 4% (w / v) paraformaldehyde, and OCT embedding agent.

[0033] (2) Cells 293T cells (for AAV packaging) were purchased from the cell bank of Peking Union Medical College and cultured in DMEM medium containing 10% (v / v) fetal bovine serum and routinely cultured in a 37°C, 5% CO2 incubator.

[0034] (3) Animals Male C57BL / 6J mice, aged 8-10 weeks and weighing 22.5g ± 2.5g, were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The animals were housed in an SPF-grade environment with a 12-hour light / 12-hour dark cycle and free access to food and water. All animal experimental procedures were approved by the animal protection and use committee of a certain institution.

[0035] (4) shRNA The short hairpin RNA targeting SAA1 (sh-SAA1) and its negative control shRNA (sh-NC) were designed and synthesized by Wuhan Shumi Brain Science Technology Co., Ltd., and cloned into AAV vectors carrying the AP2 promoter, respectively. AAV-AP2-sh-SAA1 and AAV-TBG-sh-NC were obtained and used to specifically knock down SAA1 expression in white adipose tissue.

[0036] Among them, the short hairpin RNA (sh-SAA1) targeting SAA1 is the shRNA targeting SAA1, and its sequence is as follows: The nucleotide sequence is as shown in SEQ ID NO:2, the sense strand is 5'-GCGAGCCTACACTGACATGAA-3'; the nucleotide sequence is as shown in SEQ ID NO:3, the antisense strand is 5'-TTCATGTCAGTGTAGGCTCGC-3'.

[0037] The AAV carrying the AP2 promoter was packaged and prepared by Wuhan Shumi Brain Science Technology Co., Ltd. Its capsid protein subtype is type 8, that is, the serotype is AAV8, and it is called AAV8 serotype vector or AAV-AP2 vector.

[0038] SAA1 (protein) is a human-derived SAA1 (protein) or its mammalian homolog.

[0039] The mammal in question is the mouse. The amino acid sequence of SAA1 (serum amyloid A1 from mesenteric white adipose tissue) is shown in SEQ ID NO.1: The sequence is: MKLLTGLVFCSLAVLGVSAQQWTAFISHEARQAGVQDMIRAYQDMKEANWKNSTVLVSKGNGDAAVLVPGGPEAWAAEVVSNAKENGIKAALTGRGEDSLADQAANKWGR SGKDPNRFRP KGLPDKY, where amino acids 1-18 (MKLLTGLVFC SLAVLGVSA) constitute the signal peptide sequence, and amino acids 19-122 (QQWTAFISHE... to the end) constitute the mature protein sequence. This sequence corresponds to UniProt accession number P05366.

[0040] Example I. Experimental Methods 1. Animal model establishment and grouping C57BL / 6J mice were randomly divided into a control group, a METH withdrawal model group, and an intervention group (AAV-AP2-sh-NC control group, referred to as AAV-AP2-sh-NC and AAV-AP2-sh-SAA1 intervention group, referred to as AAV-AP2-sh-SAA1).

[0041] In the METH withdrawal model group, mice were administered METH solution via intraperitoneal injection at escalating doses for 7 consecutive days, followed by discontinuation of the drug to establish a withdrawal model. The methamphetamine (METH) solution was prepared by accurately weighing an appropriate amount of methamphetamine hydrochloride, dissolving it in sterile physiological saline to prepare a METH solution of the required concentration, and then filtering it through a 0.22 μm microporous membrane for sterilization. The dosage was 2 mg / kg, administered via intraperitoneal injection once daily for 7 consecutive days, with an injection volume of 10 mL / kg body weight.

[0042] Before modeling, mice in the intervention group received in situ injections of mesenteric white adipose tissue: the mesenteric white adipose tissue was exposed by laparotomy, and AAV-AP2-sh-SAA1 and AAV-AP2-sh-NC (titer 1×10¹² vg / mL, 50 μL per mouse) were injected at multiple points. The wounds were sutured postoperatively, and METH modeling began 21 days after recovery (same as the METH withdrawal model group). The control group received an equal volume of physiological saline.

[0043] 2. Tissue sample collection At specific time points during the withdrawal period, mice in each group were euthanized after anesthesia, and peripheral major organs such as the heart and liver (including mesenteric white adipose tissue) and brain regions including the prefrontal cortex, hippocampus, and cerebellum were collected. Three mice per group were immediately cryopreserved in liquid nitrogen for RNA and protein extraction; another three mice per group had their mesenteric white adipose tissue fixed in 4% (w / v) paraformaldehyde for immunofluorescence staining.

[0044] 3. PCR detection Total RNA was extracted from the peripheral tissues and brain tissues using the TransZol Up Plus RNA Extraction Kit (catalog number: ER501) from Beijing TransGen Biotech Co., Ltd. After reverse transcription into cDNA, real-time quantitative PCR was performed using the TransStart Top Green qPCR SuperMix Kit (catalog number: AQ131) from Beijing TransGen Biotech Co., Ltd. The specific steps are as follows: ① Prepare the following reaction mixture in a 96-well qPCR plate (operate on ice): 2×TransStart Top Green qPCR SuperMix 10μL.

[0045] Forward Primer (10μM)) 0.4μL.

[0046] Reverse Primer (10μM) 0.4μL.

[0047] 1 μL of cDNA template (adjust as needed based on preliminary experimental results).

[0048] Passive Reference Dye (50×) 0.4μL.

[0049] Add RNase-free water to a final volume of 20 μL.

[0050] ② After sealing, briefly centrifuge to ensure the reaction solution is at the bottom of the well and free of air bubbles.

[0051] ③ Amplification was performed on a real-time quantitative PCR instrument. The reaction procedure is as follows: Pre-denaturation: 94℃ for 30 seconds.

[0052] PCR cycles (40 cycles): Denaturation at 94℃ for 5 seconds.

[0053] Anneal at 60℃ for 15 seconds.

[0054] Extend the exposure time to 72℃ for 10 seconds (collect fluorescence signal).

[0055] ④ Primer sequence: Target gene SAA1 The primer sequences are as follows: SAA1-F: 5'-GGAGTCTGGGCTGCTGAGAAAA-3', as shown in SEQ ID NO:4.

[0056] SAA1-R: 5'-TGTCTGTTGGCTTCCTGGTCAG-3', as shown in SEQ ID NO:5.

[0057] Internal reference gene GAPDH The primer sequences are as follows: GAPDH-F: 5'-TGTGTCCGTCGTGGATCTGA-3', as shown in SEQ ID NO:6.

[0058] GAPDH-R: 5'-TTGCTGTTGAAGTCGCAGGAG-3', as shown in SEQ ID NO:7.

[0059] Use 2 ⁻ΔΔCtThe relative expression level of SAA1 mRNA was calculated using a method with GAPDH as an internal reference gene. Each sample was tested in triplicate, and the average Ct value was used for calculation.

[0060] 4. Immunofluorescence staining White adipose tissue fixed with PFA was dehydrated, embedded in OCT, and sectioned. After blocking, the sections were incubated with rabbit-derived SAA1 primary antibody (4°C overnight), followed by incubation with fluorescently labeled secondary antibody the next day. Cell nuclei were counterstained with DAPI, mounted, and observed and photographed under a confocal microscope.

[0061] Overnight stays refer to stays of 12 hours or more.

[0062] 5. Behavioral testing The tail suspension test and forced swimming test were used to evaluate depressive-like behavior (anhedonia state) in mice, and the open field test and elevated cruciate maze test were used to evaluate anxiety-like behavior, in order to assess the METH withdrawal of negative emotions and the intervention effect. The specific procedures are as follows:

[0063] Evaluation of depressive-like behavior: In the tail suspension test, mice were suspended from a support with their tails taped 1.5 cm from the tip. Activity was recorded for 6 minutes, and the immobility time in the following 4 minutes was calculated. In the forced swimming test, mice were placed in a transparent cylindrical tank (25 cm high, 15 cm in diameter, 15 cm deep, 25°C). Activity was recorded for 6 minutes, and the immobility time in the following 4 minutes was calculated. Prolonged immobility time reflected increased depressive-like behavior.

[0064] Assessment of anxiety-like behavior: In the open field test, mice were placed in the center of a 50cm×50cm×40cm open field box, and their free movement trajectory was recorded over 10 minutes. The time spent in the central area, the number of entries, and the total distance moved were counted. In the elevated cross maze test, mice were placed on the central platform (6cm×6cm), and their movement trajectory was recorded over 5 minutes. The percentage of time spent in the open arms and the percentage of entries were counted. A shortened time spent in the central area or in the open arms reflected increased anxiety-like behavior.

[0065] All behavioral experiments were conducted in a quiet, softly lit environment, with animals acclimatizing for at least 30 minutes before testing. After each mouse test, the apparatus was cleaned with 75% (v / v) ethanol to avoid interference from residual odors. Video recordings were automatically analyzed using behavioral analysis software, and data are expressed as mean ± standard error.

[0066] II. Results 1. Specific upregulation of SAA1 expression in mesenteric white adipose tissue of METH withdrawal mice PCR and Western blot results showed that, compared with the control group, the mRNA and protein levels of SAA1 in the mesenteric white adipose tissue of mice in the METH withdrawal model group were significantly upregulated, while the expression level of SAA1 in liver tissue showed no significant change (see...). Figure 1 A and Figure 1 (B in the middle).

[0067] The above results indicate that the increase of SAA1 during METH withdrawal is specific to peripheral adipose tissue.

[0068] 2. Immunofluorescence staining to verify the SAA1 upregulation trend Immunofluorescence staining of mesenteric white adipose tissue revealed that the fluorescence intensity of SAA1 in the adipose tissue of the METH withdrawal model group mice was significantly higher than that in the control group, further confirming the upregulation trend of SAA1 protein in adipose tissue (see [link to relevant documentation]). Figure 1 (C in the middle).

[0069] 3. Knockdown of SAA1 in white adipose tissue can alleviate elevated peripheral SAA1 levels and improve negative behaviors. By injecting AAV-TBG-sh-SAA1 into mesenteric fat in situ, and utilizing the TBG promoter to drive the specific expression of shRNA in white adipose tissue, targeted knockdown of SAA1 in mesenteric white adipose tissue was successfully achieved (see the overall procedure). Figure 2 (A in the text). The results showed that, compared with the AAV-AP2-sh-NC control group, the level of SAA1 in the peripheral plasma of mice in the AAV-AP2-sh-SAA1 intervention group was significantly decreased (see A). Figure 2 B and Figure 2 (C in the text). Meanwhile, behavioral tests showed that mice in the sh-SAA1 intervention group spent significantly more time in the central area of ​​the open field and entered the elevated cruciform maze more frequently (suggesting a reduction in anxiety-like behavior, see [reference]). Figure 2 D in Figure 2 E and Figure 2 In the F section, the immobility time was significantly reduced in tail suspension and forced swimming behaviors (suggesting that anorexic stimulation was alleviated, see F). Figure 2 G and Figure 2 (H in the text).

[0070] The above results indicate that targeting SAA1 in mesenteric white adipose tissue can effectively alleviate negative emotions caused by METH withdrawal.

[0071] Based on the above experimental results, the feasibility and superiority of the SAA1 inhibitor provided by this invention in the preparation of drugs for treating METH addiction withdrawal syndrome and preventing relapse have been fully verified. It can effectively overcome the limitations of existing clinical treatments described in the background section, as detailed below: (1) Precise intervention targeting the etiology solves the problem of "target mismatch" of existing drugs. Existing conventional psychiatric drugs (such as SSRIs, antidepressants) mainly act on the monoamine neurotransmitter system, and their pharmacological mechanisms are "misaligned" with the specific pathological processes driven by METH neurotoxicity, thus their efficacy is uncertain. The core of the technical solution of this invention lies in precisely targeting the SAA1-SCARB1 pathway, a newly revealed core mechanism of METH withdrawal.

[0072] This invention is the first to demonstrate that SAA1 derived from peripheral mesenteric white adipose tissue is a key molecule driving negative emotions during METH withdrawal. By using an SAA1 inhibitor (specifically, AAV-AP2-shSAA1 for SAA1 knockdown in this invention), the signaling source of the pathological pathway "peripheral adipose tissue-medial prefrontal cortex" was directly blocked. PCR and immunofluorescence results consistently showed that this intervention strategy effectively inhibited the abnormal upregulation of SAA1 in mesenteric white adipose tissue, proving that it achieved precise intervention from the molecular target to the tissue source, fundamentally solving the problem of target misalignment in existing drugs that only treat the symptoms and not the root cause.

[0073] (2) Intervening in the peripheral source and avoiding central side effects solved the problem of "poor compliance". Existing psychiatric medications often lead to poor patient adherence due to central nervous system side effects such as drowsiness and cognitive impairment. The technical solution of this invention has the unique advantage of prioritizing peripheral intervention. SAA1 inhibitors (such as the adipose tissue-specific knockdown strategy used in this embodiment) primarily act on peripheral white adipose tissue, indirectly regulating central nervous system function by reducing peripheral SAA1 levels, rather than directly acting on the central neurotransmitter system.

[0074] This "peripheral intervention of the central nervous system" strategy can theoretically avoid the widespread central nervous system side effects caused by traditional drugs acting directly on the whole brain. Behavioral results showed that mice receiving AAV-AP2-shSAA1 intervention experienced improvement in negative emotions without significant abnormalities in motor function or general condition, suggesting that this strategy has good safety potential and may solve the problem of low adherence caused by the side effects of existing treatments.

[0075] (3) It blocks the key drivers of relapse and solves the problem of "inability to prevent relapse". Negative emotions during withdrawal are a high-risk factor for relapse, and current treatments lack effective means to prevent relapse. The technical solution of this invention directly alleviates negative emotions, cutting off a key driving factor for relapse at its source.

[0076] Behavioral results confirmed that specific knockdown of SAA1 significantly improved depressive-like behaviors (forced swimming and tail-hanging behaviors) and anxiety-like behaviors (increased time spent in the center of the open field and increased number of times the arms were opened in the elevated cruciate maze) in METH withdrawal mice. The relief of negative emotions indicates a reduced motivation for individuals to seek medication to alleviate emotional distress, thereby effectively reducing the risk of relapse. This suggests that SAA1 intervention not only treats acute withdrawal symptoms but also has potential application value in preventing relapse, filling a gap in existing technologies in this field.

[0077] In summary, this invention, by precisely targeting the newly discovered "SAA1-fat-brain axis" pathogenesis mechanism, effectively intervenes in the negative emotions associated with METH withdrawal from a peripheral source, overcoming the shortcomings of existing drugs such as target mismatch, significant side effects, and inability to prevent relapse. Therefore, the technical solution protected by this invention truly solves the technical problems existing in the prior art as described in the background section, and has significant clinical translational potential and application value.

[0078] It should be noted that when numerical ranges are involved in this invention, it should be understood that the two endpoints of each numerical range and any value between the two endpoints can be selected. To avoid redundancy, this invention describes preferred embodiments.

[0079] Although preferred embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments, all of which fall within the scope of the invention.

Claims

1. Application of SAA1 protein inhibitors in the preparation of drugs for treating methamphetamine addiction withdrawal syndrome.

2. The application according to claim 1, characterized in that, The SAA1 protein is a human SAA1 protein or its mammalian homolog.

3. The application according to claim 2, characterized in that, The SAA1 protein inhibitor is at least one of the following: Antibodies or antigen-binding fragments thereof that specifically bind to the SAA1 protein; Small molecule inhibitors of SAA1 protein; siRNA, shRNA, antisense oligonucleotides, or miRNAs that target the SAA1 protein; A CRISPR / Cas9 gene editing system targeting the gene encoding the SAA1 protein; A carrier system for specifically delivering the SAA1 protein inhibitor to mesenteric white adipose tissue.

4. The application according to claim 3, characterized in that, The vector system is an adeno-associated virus vector, which contains a white adipose tissue-specific promoter. The white adipose tissue-specific promoter is either the mouse AP2 promoter or the human FABP4 promoter.

5. The application according to claim 4, characterized in that, The adeno-associated virus vector is the AAV8 serotype vector.

6. The application according to claim 3, characterized in that, The shRNA of the mRNA targeting the SAA1 protein comprises a sense strand with a nucleotide sequence as shown in SEQ ID NO:2 and an antisense strand with a nucleotide sequence as shown in SEQ ID NO:

3.

7. The application according to claim 3, characterized in that, The antibody is a monoclonal antibody or a polyclonal antibody; The monoclonal antibody is selected from humanized antibodies, murine monoclonal antibodies, or chimeric antibodies.

8. The application according to any one of claims 1 to 7, characterized in that, The drug is a liquid preparation.

9. The application according to any one of claims 1 to 7, characterized in that, The drug is used for local or systemic administration to the mesenteric white adipose tissue.

10. The application according to any one of claims 1 to 7, characterized in that, The methamphetamine addiction withdrawal syndrome includes anxiety, depression, irritability, or other negative emotional symptoms that occur during withdrawal.

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