Application of UXS1 as biomarker or UXS1 detection substance and pharmaceutical composition

Abstract

The invention provides application of UXS1 as a biomarker or a substance for detecting UXS1 and a pharmaceutical composition, and the application is as follows: application in preparation and / or screening of products for diagnosis or auxiliary diagnosis of metformin treatment sensitivity of lung adenocarcinoma patients; application in preparing and / or screening a product for monitoring the curative effect of a lung adenocarcinoma patient; the invention also relates to application in preparing and / or screening products for evaluating the metformin resistance of lung adenocarcinoma patients. The technical scheme provided by the invention has the following advantages: 1, the specific molecular mechanism that metformin releases allosteric inhibition by inducing UGDH S476 site phosphorylation is clarified; 2, a metabolic regulation network for inhibiting and triggering UDP-uronic acid toxicity accumulation by the UXS1 function is disclosed; 3, a synergistic treatment scheme for enhancing the lethal effect of UXS1 inhibitor synthesis through metformin pretreatment is provided; and 4, establishing an immune collaborative treatment scheme based on uronic acid metabolism reprogramming.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of UXS1 as a biomarker or a substance for detecting UXS1, as well as pharmaceutical compositions. Background Technology

[0002] According to Global Cancer Statistics 2024 and the "2022 China Cancer Incidence and Mortality Report" released by the National Cancer Center of China in 2024, lung cancer ranks first in both incidence and mortality among all malignant tumors globally and in my country, seriously threatening public health. Lung adenocarcinoma, as the most prevalent pathological subtype of lung cancer, accounts for nearly two-thirds of all new lung cancer cases each year. Lung adenocarcinoma is metabolically active, proliferates rapidly, and is prone to recurrence and metastasis, resulting in unsatisfactory treatment outcomes and prognosis for patients, with an overall 5-year survival rate of only about 30%.

[0003] Therefore, further exploration of the metabolic characteristics of lung adenocarcinoma is of great significance for finding more precise and effective treatment options, reducing recurrence and metastasis, and thus improving patient prognosis. Metformin research presents contradictions: in vitro experiments have confirmed that it can inhibit tumor cell proliferation through pathways such as AMPK / mTOR, but clinical cohort studies have shown no statistically significant improvement in the survival of lung adenocarcinoma patients.

[0004] A recent study reported a "pool model" of UGDH / UXS1—UGDH acts as a "faucet," being highly expressed in cancer cells and thus promoting the production of UDPGA. Excessive UDPGA can disrupt the morphology and function of the Golgi apparatus, "killing" cancer cells; while UXS1, a metabolic enzyme located on the Golgi apparatus, acts as a "drainage pipe," converting toxic UDPGA into non-toxic UDP-xylose, thereby preventing the accumulation of UDPGA and its damage to cells.

[0005] There are currently no reported small molecule inhibitors targeting UXS1.

[0006] Tumor immunology is an important area of ​​development in cancer treatment, where the body uses immune responses to recognize, respond to, and eliminate tumor cells. However, during tumor development, tumor cells adapt to changes in the immune microenvironment through various means and ultimately overcome the immune system's attack, thus achieving continuous growth.

[0007] According to literature reports, metabolites such as UDPG and UDPGA in the uronic acid pathway are closely related to immunity. UDPG is a natural agonist of the G protein-coupled P2Y14 receptor, present in the immune system, promoting neutrophil recruitment and leading to the release of pro-inflammatory chemokines. Accumulation of UDPGA significantly alters the function and morphology of the Golgi apparatus, ultimately inducing tumor cell apoptosis. The antigens released after tumor cell apoptosis can activate the body's immune system, triggering a specific immune response. Therefore, targeting UXS1 in combination with metformin may have a significant impact on the immune microenvironment through metabolites in the uronic acid pathway. Summary of the Invention

[0008] In view of the deficiencies in the prior art, the object of the present invention is to provide the application of UXS1 as a biomarker or a substance for detecting UXS1, as well as pharmaceutical compositions.

[0009] To achieve the above objectives, the present invention proposes the following technical solution: In a first aspect, the present invention proposes the use of UXS1 as a biomarker or a substance for detecting UXS1 in at least one of the following: Application in the preparation and / or screening of products for the diagnosis or auxiliary diagnosis of metformin treatment sensitivity in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for monitoring the efficacy of treatment in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for the evaluation of metformin resistance in patients with lung adenocarcinoma.

[0010] Preferably, the substance for detecting UXS1 is selected from substances for detecting UXS1 protein or its mRNA, and the substance for detecting UXS1 protein or its mRNA includes any reagent required to detect the expression level of UXS1 protein or mRNA in tumor tissue by immunohistochemistry or qRT-PCR.

[0011] Secondly, the present invention proposes the use of UXS1 as a target or UXS1 inhibitor in the preparation and / or screening of drugs for treating patients with lung adenocarcinoma who are sensitive to metformin treatment. The UXS1 inhibitor includes substances that reduce the expression level, content or activity of UXS1 protein, or substances that inhibit the expression of the UXS1 gene.

[0012] Preferably, the UXS1 inhibitor is psyllium glycoside D or a pharmaceutically acceptable salt or structurally modified derivative thereof.

[0013] Preferably, the structural modification derivative of the UXS1 inhibitor is an acetylated, glycosylated, or sulfonated derivative of psyllium glycoside.

[0014] In the technical solution described in this invention, the mechanism of action of the drug is as follows: Metformin activates PKCζ kinase via an AMPK-independent pathway, catalyzing phosphorylation of UGDH S476. Phosphorylation of UGDH relieves its allosteric inhibition and promotes the synthesis of UDP-glucuronic acid; UXS1 inhibition prevents UDP-uronic acid from being converted to UDP-xylose, inducing endoplasmic reticulum stress and mitochondrial apoptosis; The accumulation of metabolites activates tumor immunity, promotes CD8+ T cell infiltration and IFN-γ secretion, and promotes the transformation of macrophages into M1 cells.

[0015] Thirdly, the present invention provides a method for screening pharmaceutical compositions that are sensitive to metformin treatment in patients with lung adenocarcinoma, the method comprising the following steps: The target was determined based on molecular mechanisms and regulatory pathways, and the target was UXS1. Candidate compounds were obtained through virtual screening of a compound library and experimental screening. The candidate compound was used in combination with metformin to determine the final drug composition.

[0016] Fourthly, this invention proposes the application of an UGDH sulfation level regulating agent in the preparation and / or screening of metformin for the treatment of patients with lung adenocarcinoma, wherein the UGDH sulfation level regulating agent is an agent that reduces the sulfation modification sites of UGDHS476.

[0017] Fifthly, the present invention provides a pharmaceutical composition for treating metformin-sensitive lung adenocarcinoma patients, the pharmaceutical composition comprising an effective dose of metformin, the aforementioned UXS1 inhibitor, and / or the aforementioned UGDH sulfation level modulator.

[0018] Preferably, the molar ratio of metformin (1mM) to the UXS1 inhibitor is 1:0.01 to 1:5, more preferably 1:0.1 to 1:2.

[0019] Preferably, the effective dose of metformin is 0.5-2 g / day; and the effective dose of the UXS1 inhibitor psyllium glycoside D is 0.1-10 mg / kg / day.

[0020] Sixthly, the present invention proposes the use of the above-described pharmaceutical composition in at least one of the following: Application in the preparation and / or screening of products for the diagnosis or auxiliary diagnosis of metformin treatment sensitivity in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for monitoring the efficacy of treatment in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for the evaluation of metformin resistance in patients with lung adenocarcinoma.

[0021] In the technical solution described in this invention, the applicant attempted to find small molecule inhibitors targeting UXS1 through virtual screening of a compound library and preliminary experimental verification. Through virtual screening of a compound library consisting of 17,676 small molecules, the applicant preliminarily identified 126 compound inhibitors that may bind to UXS1.

[0022] Further experimental verification revealed that the combination of plantanoside D and metformin exhibited a significant synthetic lethal effect.

[0023] Compared with the prior art, the present invention has the following advantages and beneficial effects: 1. Elucidate the specific molecular mechanism by which metformin relieves allosteric inhibition by inducing phosphorylation at the UGDH S476 site. 2. Revealing the metabolic regulatory network of UDP-glucuronic acid toxicity accumulation caused by UXS1 functional inhibition. 3. To provide a synergistic treatment regimen that enhances the synthetic lethal effect of UXS1 inhibitors through metformin pretreatment. 4. Establish an immune synergistic therapy based on uronic acid metabolism reprogramming. Attached Figure Description

[0024] Figure 1 The combined use of UXS1 and metformin produced a synergistic lethal effect: A shows the three-dimensional structure of UXS1 and the two-dimensional structure of the target, with the three-dimensional structure of the target highlighted in red; B shows the sensitizing effect of the targeted compound obtained at a concentration of 10 μM on A549 and PC9 cells in combination with metformin, compared to the DMSO control; C shows the IC50 of three UXS1 inhibitors combined with 1 mM metformin on A549 and PC9 cells as determined by the CCK8 assay, where the three UXS1 inhibitors are proctolin, plantanoside, and N-feruloyloctopamine; D shows plantanoside... Two-dimensional and three-dimensional structures; E shows the comparison of cell apoptosis after different concentrations of plantago asiatica combined with different concentrations of metformin; F shows the effect of plantago asiatica combined with 1mM metformin on the proliferation of A549 and PC9 cells in the cell proliferation experiment, where "-" indicates no addition and "+" indicates addition; G shows the organoid model verification that the combination of plantago asiatica and metformin produces the strongest anti-tumor effect; H shows the CDX model results, which verify that the combination of plantago asiatica and metformin produces the strongest anti-tumor effect; I shows the tumor growth curve of the CDX model; K shows the spontaneous lung cancer model showing that the group using the combination of plantago asiatica and metformin has the longest survival time; J shows that regular CT follow-up confirms that the group using the combination of plantago asiatica and metformin has the slowest tumor growth.

[0025] Figure 2 This study demonstrated the effects of UXS1 targeting combined with metformin on the immune microenvironment of LUAD (Low-Induced Luteinized Adenoiditis). Specifically, A showed the difference in the association between tumor cells and immune cells in LUAD patients treated with metformin and those with high / low UXS1 expression; B showed the association between T cell (CD8) and M2-TAM (CD68, CD163) infiltration in tumor tissues of LUAD patients treated with metformin and those with high / low UXS1 expression, as shown by immunohistochemistry and immunofluorescence; C showed the expression of CD8 and IFNγ in T cells and macrophage differentiation (M1-TAM: CD86, CD68; M2-TAM: CD206, CD163) after co-culturing LUAD cells by flow cytometry, with the addition of metformin or plantago asiatica glycoside, or a combination of both; D showed the expression of CD8 and IFNγ in T cells and macrophage differentiation (M1-TAM: CD86, CD68; M2-TAM: CD206, CD163) after co-culturing LUAD cells by qRT-PCR. The treatment options include the addition of metformin or plantagoside, or a combination of metformin and plantagoside. Flowchart E illustrates the experimental design of the spontaneous lung cancer model, including four treatment groups: immunotherapy alone, immunotherapy + metformin, immunotherapy + plantagoside, and immunotherapy + metformin + plantagoside, detailing model establishment, treatment timelines, and sample collection. Kaplan-Meier survival curves compare the survival times of the four treatment groups, where Immunothreapy represents immunotherapy alone, Immunothreapy + Metformin represents immunotherapy + metformin, and Immunothreapy + Plantagoside represents immunotherapy + plantagoside. hreapy+Plantainoside+Metfromin represents immunotherapy plus plantago asiatica glycoside; G shows computed tomography (CT) scans assessing tumor growth dynamics in the four treatment groups, where Immunothreapy represents immunotherapy alone, Immunothreapy+Metfromin represents immunotherapy plus metformin, Immunothreapy+Plantainoside represents immunotherapy plus plantago asiatica glycoside, and Immunothreapy+Plantainoside+Metfromin represents immunotherapy plus plantago asiatica glycoside; H shows representative histological (HE) and immunofluorescence staining results of lung tissue from mice in the four treatment groups, with M2-targeted tumor-associated macrophages and CD8+ cells shown in the figure. + IFNγ + T cells. Detailed Implementation

[0026] The present invention will now be described in detail with reference to specific embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that those skilled in the art can make several adjustments and improvements without departing from the concept of the present invention. These all fall within the scope of protection of the present invention.

[0027] The experimental design concept of this patent application is as follows: 1. Mechanism studies revealed that metformin relieves allosteric inhibition by inducing phosphorylation at the UGDH S476 site, as detailed below: Metformin activates the AMPK signaling pathway by inhibiting mitochondrial respiratory chain complex I, leading to an increased AMP / ATP ratio. Activated AMPK directly phosphorylates the S476 site of UGDH (UDP-glucose dehydrogenase), relieving its allosteric inhibition and thereby enhancing UGDH enzyme activity and promoting the production of UDP-glucuronic acid (UDPGA). This process can be verified by co-immunoprecipitation (Co-IP) and mass spectrometry analysis, and the relief of allosteric inhibition can be confirmed by in vitro enzyme activity assays.

[0028] 2. A metabolic regulatory network was discovered that leads to the accumulation of UDP-glucuronic acid toxicity due to UXS1 functional inhibition, as detailed below: UXS1 is a key enzyme for clearing UDPGA. Its inhibition leads to the accumulation of UDPGA in the Golgi apparatus, disrupting the morphology and function of the Golgi apparatus.

[0029] The toxicity of UDPGA is achieved through the following pathways: Golgi apparatus dysfunction: inhibits surface receptor transport and reduces cell signal transduction capacity.

[0030] Metabolic reprogramming: UDPGA accumulation feedback inhibits UGDH activity, forming a negative regulatory loop.

[0031] UDPGA levels can be detected by metabolomics (such as UPLC-MS), and changes in Golgi apparatus structure can be observed by immunofluorescence.

[0032] 3. A synergistic treatment regimen that enhances the synthetic lethal effect of UXS1 inhibitors through metformin pretreatment; Metformin increases the supply of UDPGA substrates by upregulating UGDH expression and activity, making cancer cells more dependent on UXS1 for detoxification. Combining this with UXS1 inhibitors (such as psyllidin) can lead to a significant increase in UDPGA toxicity, triggering a synthetic lethal effect.

[0033] Therefore, the experimental design includes the following: Cell models: A549 and PC9 cells were pretreated with metformin and then UXS1 inhibitor was added. Changes in IC50 were detected by CCK8 assay.

[0034] Animal models: CDX model to verify the tumor-suppressive effect of combination therapy, and synergistic effect was analyzed by tumor growth curve and survival period.

[0035] 4. Establish an immune synergistic therapy regimen based on uronic acid metabolism reprogramming, which includes: UDPGA accumulation can reshape the tumor immune microenvironment. UPLC-MS was used to quantify UDP-glucuronide levels to confirm its accumulation after UXS1 inhibition. This process involves endoplasmic reticulum stress and apoptosis; therefore, the effects were determined by detecting endoplasmic reticulum stress markers such as PERK and CHOP, as well as mitochondrial apoptosis pathways (e.g., Caspase-3 activation). Furthermore, the accumulation of metabolites activates tumor immunity, promotes CD8+ T cell infiltration and IFN-γ secretion, and promotes macrophage transformation into M1 cells.

[0036] Immune cell infiltration: Specifically, high UDP levels are used to promote the infiltration of M2 macrophages (CD68+, CD163+) and CD8+ T cells; Combination immunotherapy: The regimen uses metformin, a UXS1 inhibitor, and a PD-1 antibody in combination to enhance the anti-tumor immune response. Furthermore, flow cytometry was used to detect CD8+ T cell and IFNγ expression, and the survival-prolonging effect was verified using a spontaneous lung cancer model.

[0037] Example 1 In this embodiment, the pharmaceutical composition is metformin combined with an UXS1 inhibitor (e.g., psyllium glycoside D).

[0038] In this embodiment, the combined application method employs sequential drug delivery, as detailed below: a) Administer metformin to the patient at a dose ranging from 0.5 to 2 g / day; b) The subsequent use of the UXS1 inhibitor plantagoside D at a dose range of 0.1-10 mg / kg / day induced UDP-glucuronic acid accumulation.

[0039] Further treatment will be administered in combination with immune checkpoint inhibitors based on the patient's PD-1 / PD-L1 expression status.

[0040] In the above embodiments, the UXS1 inhibitor is administered orally, and the administration time window is within 2 hours after metformin administration.

[0041] In addition, the inventors discovered that UXS1 inhibition prevents UDP-uronic acid from being converted into UDP-xylose, inducing endoplasmic reticulum stress and mitochondrial apoptosis.

[0042] The collaborative treatment plan is designed as follows: Cell model: Based on A549 / PC9 cells, the changes in IC50 of UXS1 inhibitors (such as psyllium glycoside) after metformin pretreatment were compared to verify the synergistic effect.

[0043] Animal models: According to the CDX model, the tumor volume and survival time in the combination therapy group were significantly better than those in the single therapy group.

[0044] Immune microenvironment analysis: The proportion of CD8+ T cells and M1 macrophages (CD86+) was detected by flow cytometry, while IFN-γ expression was measured by qRT-PCR.

[0045] Combined immunotherapy: In spontaneous lung cancer models, triple therapy (immunotherapy + metformin + psyllium glycoside) significantly prolonged survival.

[0046] Example 2: Determination of metformin and psyllium glycoside concentrations (related results can be found in...) Figure 1 (C and D) In this embodiment, CCK8 and EdU experiments were performed, and the results consistently showed that the combination of plantago asiatica glycoside and metformin significantly enhanced the cytotoxicity against tumor cells (see [link to relevant documentation]). Figure 1 C and D, by Figure 1 The optimal concentration was found to be 1 mM metformin and 1 μM plantago asiatica glycoside. A ZIP regression model (which integrates the Bliss and Loewe models and assumes no interaction) was used to assess the synergistic inhibitory effect of metformin and plantago asiatica glycoside. The results further confirmed the significant synergistic effect of the combination (see results below). Figure 1 E).

[0047] Example 3: Validation of the in vitro experimental system The validation results of the aforementioned in vitro experimental system can be found in [reference needed]. Figure 1 E and F.

[0048] Depend on Figure 1 As can be seen from E and F, in this embodiment, the synthetic lethal effect of the UXS1-targeting inhibitor and metformin on lung adenocarcinoma cells was detected, and the experimental results are as follows: 1. In vitro detection of the binding interaction between plantago asiatica glycoside D and UXS1 (using the Taoshu SPR / MST assay). 2. To verify the regulation of the uronic acid pathway by plantago asiatica glycoside D and its synergistic effect with metformin in lung adenocarcinoma cells, and to compare it with the currently reported UGDH inhibitor 4-MU. 3. Plantago asiatica glycoside D was used in combination with metformin to treat UGDH-KO, UGDHS476A, UGDHS476D and UXS1-KO cells respectively, which formally confirmed that plantago asiatica glycoside D exerts its effect by inhibiting UXS1.

[0049] In addition, the synergistic effect of UXS1-targeting inhibitors and metformin on co-cultured T cells and macrophages was investigated, and the experimental results are as follows: 1. To investigate the effect of adding a small molecule compound targeting UXS1 and / or metformin to lung adenocarcinoma cells and co-culturing them with T cells and macrophages on the content of uronic acid pathway metabolites in the supernatant of immune cells.

[0050] 2. The role of detecting biomarkers and functions of co-cultured T cells and macrophages; 3. To detect the effects of small molecule compounds targeting UXS1 and / or metformin itself on T cell and macrophage-related markers and functions.

[0051] The above results show that metformin combined with plantain glycoside D significantly promotes immunity, significantly increases the number of CD8+ T cells infiltrating tumors, significantly increases the number of M1 macrophages, and significantly decreases the number of M2 macrophages.

[0052] Example 4: Validation of Organoid Model In this embodiment, the experimental procedure is as follows: A portion of the tumor tissue and a section of unaffected lung tissue (0.5-2 cm³) were taken for analysis to confirm the presence of non-small cell lung cancer (NSCLC). The remaining tissue samples were transferred to liquid culture medium containing Hank's buffer and antibiotics (catalog number 15240096, ThermoFisher). The tumor tissue samples were cut into small pieces, digested, and digested using type II collagenase and DNase I at 37°C to obtain a mixture; The mixture was continuously stirred at 120 rpm, and trypsin was added for digestion for 10 minutes. The mixture was then passed through a 70 µm filter to remove any large pieces of tissue, and the desired cells were obtained after filtration.

[0053] Gently mix the desired cells with the matrix gel and place 30–50 µL droplets into a 6-well plate that has been heated to the appropriate temperature. Then invert the droplets for one minute, followed by 10 minutes in an incubator to allow them to solidify.

[0054] Add 2.5 mL of warm organoid culture medium to each well, and fill any empty wells with sterile phosphate-buffered saline (PBS) to prevent culture medium evaporation.

[0055] Organoids were cultured until they reached a diameter of 100-150 µm over a two-week period. The resulting samples were then analyzed; the results are detailed below. Figure 1 G.

[0056] In addition, a nude mouse subcutaneous xenograft model was constructed. Combining the results of the organoid model and the nude mouse subcutaneous xenograft model, it can be seen that the combination drug regimen described in this invention shows a significant synergistic inhibitory effect on the growth of both organoids and subcutaneous tumors.

[0057] Example 5: Validation of an animal model of in situ lung tumor formation (see relevant experimental results) Figure 1 JK) In this embodiment, the animal model of in situ lung tumorigenesis was constructed using the following steps: An expression vector containing the humanized UGDH gene was constructed. Using CRISPR / Cas9 gene editing technology, the UGDH gene in LLC cells (Lewis lung cancer cells) derived from the lungs of C57BL / 6 mice was knocked out and then transformed into humanized UGDH.

[0058] A mouse model of in situ lung cancer tumorigenesis was established by injecting humanized UGDH LLC cells via the tail vein (i.e., obtaining an animal model of in situ lung cancer tumorigenesis).

[0059] Subsequently, patients were treated with metformin, the optimal UXS1 inhibitor (i.e., plantain glycoside), metformin + the optimal UXS1 inhibitor plantain glycoside, and saline. Tumor size and tumor characteristics were measured, and it was found that the tumors were smallest in the combination drug group.

[0060] In addition, long-term observation of mice that spontaneously formed tumors as described above was conducted using CT imaging; the results are shown in [link to results]. Figure 1 J~K.

[0061] Combination Figure 1 As can be seen from J~K, the combination therapy group using the combination drug regimen described in this invention (in this treatment group, the dose range of metformin is 50-2000 mg / m² / day, and the dose range of the UXS1 inhibitor is psyllium glycoside D, which is 0.1-10 mg / kg / day; in addition, the immunotherapy group uses tislelizumab for immunotherapy, with a dose range of 3 mg / kg / day) had the longest survival time, and its tumor growth rate was significantly slower than that of the metformin or psyllium glycoside monotherapy groups, further confirming the synergistic lethal effect of the combination of metformin and psyllium glycoside.

[0062] In the aforementioned mouse tumors, the number of CD8+ T cells, M1, and M2 macrophages infiltrating the tumors was determined by multicolor immunofluorescence.

[0063] Furthermore, mouse tumor tissue samples were mechanically disrupted and enzymatically digested to release immune cells. Flow cytometry and magnetic bead sorting techniques were used to separate and purify the immune cells, obtaining specific types of immune cell populations. Flow cytometry was then used to perform multi-parameter analysis of the immune cells, including cell surface markers and intracellular cytokines. Combined with data analysis software, the composition, proportion, and function of the immune cells were quantitatively assessed.

[0064] A mouse model of primary lung cancer was established. The growth curves of mouse tumors after anti-PD-1 treatment, anti-PD-1 + metformin, and anti-PD-1 + metformin + UXS1 inhibition treatment were compared. It was found that the tumors in the three-drug combination group grew the slowest.

[0065] Example 6: Pharmaceutical composition for the treatment of lung adenocarcinoma In this embodiment, the pharmaceutical composition comprises a therapeutically effective amount of metformin and an UXS1 inhibitor, wherein the UXS1 inhibitor is selected from psyllium glycoside D, its pharmaceutically acceptable salt, or structurally modified derivatives. The metformin dosage range is 0.5-2 g / day, and the psyllium glycoside D dosage range is 0.1-10 mg / kg / day. Immunotherapy is strictly performed according to the lung adenocarcinoma treatment guidelines, using tislelizumab at a dose of 300 mg per cycle.

[0066] Example 7: Combined drug administration method for treating lung adenocarcinoma In this embodiment, the combined administration method includes the following steps: Metformin was administered to the patient at a dose ranging from 0.5 to 2 g per day. The UXS1 inhibitor plantagoside D was administered at a dose ranging from 0.1 to 10 mg / kg / day.

[0067] In the above embodiments, the UXS1 inhibitor is administered orally, and the administration time window is within 2 hours after metformin administration.

[0068] Example 8: Application of the pharmaceutical composition in the preparation of drugs that enhance antitumor immune responses (experimental results are shown in [reference]). Figure 2 (C and D) A co-culture system of tumor cells with T cells and macrophages was established, and it was found that the combination of metformin and psyllidine had a synergistic effect on LUAD cells co-cultured with T cells and M0 macrophages. After 48 hours of co-culture, compared with the metformin monotherapy group, the CD8 expression of T cells in the metformin and psyllidine combination group was significantly increased. This group also showed a significant decrease in M2 cells and a significant increase in M1 cells. Figure 2 (C).

[0069] Furthermore, in this embodiment, the expression of CD8 and IFNγ in T cells under co-culture conditions was detected by flow cytometry. The results showed that the proportion of CD8 and IFNγ double-positive T cells was highest in the group receiving both metformin and psyllidine treatment. Figure 2 (D).

[0070] By activating the immune system, the infiltration of CD8+ T cells and M1 macrophages in the tumor microenvironment is promoted, and the infiltration rate is increased compared with monotherapy.

[0071] Specific embodiments of the present invention have been described above. It should be understood that the present invention is not limited to the specific embodiments described above, and those skilled in the art can make various modifications or variations within the scope of the claims.

Claims

1. The use of UXS1 as a biomarker or a substance for detecting UXS1 in at least one of the following: Application in the preparation and / or screening of products for the diagnosis or auxiliary diagnosis of metformin treatment sensitivity in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for monitoring the efficacy of treatment in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for the evaluation of metformin resistance in patients with lung adenocarcinoma.

2. The application according to claim 1, characterized in that, The substance used to detect UXS1 is selected from substances that detect UXS1 protein or its mRNA, and the substance used to detect UXS1 protein or its mRNA includes any reagents required to detect the expression level of UXS1 protein or mRNA in tumor tissue by immunohistochemistry or qRT-PCR.

3. The use of UXS1 as a target or UXS1 inhibitor in the preparation and / or screening of drugs for treating patients with lung adenocarcinoma who are sensitive to metformin, wherein the UXS1 inhibitor includes substances that reduce the expression level, content, or activity of UXS1 protein, or substances that inhibit the expression of the UXS1 gene.

4. The application according to claim 3, characterized in that, The UXS1 inhibitor is psyllium glycoside D or its pharmaceutically acceptable salt or structurally modified derivative.

5. The application according to claim 4, characterized in that, The structurally modified derivatives of the UXS1 inhibitor are acetylated, glycosylated, or sulfonated derivatives of plantago asiatica glycoside.

6. A method for screening pharmaceutical compositions that are sensitive to metformin in treating patients with lung adenocarcinoma, characterized in that, The method includes the following steps: The target was determined based on molecular mechanisms and regulatory pathways, and the target was UXS1. Candidate compounds were obtained through virtual screening of a compound library and experimental screening. The candidate compound was used in combination with metformin to determine the final drug composition.

7. The use of an UGDH sulfation level regulating agent in the preparation and / or screening of metformin for the treatment of patients with lung adenocarcinoma, wherein the UGDH sulfation level regulating agent is an agent that reduces the sulfation modification sites of UGDHS476.

8. A pharmaceutical composition for treating metformin-sensitive patients with lung adenocarcinoma, characterized in that, The pharmaceutical composition comprises an effective dose of metformin, an UXS1 inhibitor as described in any one of claims 3-5, and / or an UGDH sulfation level modulator as described in claim 6.

9. The pharmaceutical composition according to claim 8, characterized in that, The effective dose of metformin is 0.5-2 g / day; the effective dose of the UXS1 inhibitor plantagoside D is 0.1-10 mg / kg / day.

10. The use of the pharmaceutical composition of claim 9 in at least one of the following: Application in the preparation and / or screening of products for the diagnosis or auxiliary diagnosis of metformin treatment sensitivity in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for monitoring the efficacy of treatment in patients with lung adenocarcinoma; Application in the preparation and / or screening of products for the evaluation of metformin resistance in patients with lung adenocarcinoma.

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