A pharmaceutical composition for treating glioblastoma

Abstract

The present invention discloses a pharmaceutical composition for treating brain glioblastoma, belonging to the field of biomedicine technology. The present invention proves through a large number of experiments that the pharmaceutical composition, including one or more of Megasphaerococcus elsdenii, Bacteroides monomorpha, Bacteroides acidogenicus and Bacteroides fragilis, effectively plays an auxiliary therapeutic role on glioblastoma.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and particularly relates to a pharmaceutical composition for treating glioblastoma. Background Art

[0002] Glioblastoma Multiforme (GBM) is the most common malignant central nervous system tumor and is difficult to treat due to its high invasiveness and rapid progression. Despite advances in traditional treatments such as surgery, radiotherapy, and chemotherapy in recent years, the median survival time of glioblastoma patients is still less than 15 months, and the five-year survival rate is extremely low. The limitations of traditional treatments are mainly due to the high heterogeneity of tumors and their resistance to treatment, which has prompted researchers to explore new treatment strategies.

[0003] Recent studies have shown that the gut microbiota may play a regulatory role in central nervous system diseases, especially the interaction of the gut-brain axis has been shown to affect the pathological processes of the central nervous system. In particular, the gut microbiota may affect the growth of brain tumors and the patient's ability to respond to treatment through its metabolites (such as short-chain fatty acids) and immune regulation. For example, some studies have found that changes in specific intestinal strains are associated with the survival time of glioblastoma patients, suggesting that intestinal microorganisms may affect tumor growth by regulating systemic responses and immune regulation. In addition, intervention strategies targeting the gut microbiota, such as the use of probiotics, are also considered as possible treatments.

[0004] Probiotics such as Lactobacillus spp. Lactobacillus ) and Bifidobacterium spp. ( Bifidobacterium ) have shown the potential to inhibit tumor growth in animal models, which may be achieved by enhancing the tumor surveillance function of the immune system and regulating inflammation. These findings provide a new perspective for the treatment of glioblastoma, which may change the current treatment paradigm and improve patient prognosis. Therefore, a deeper understanding of how the gut-brain axis affects the development of glioblastoma and how to use the intestinal microbiota for therapeutic intervention has become a hot topic of research.

[0005] However, there are no reports on the potential application of the four microorganisms, Megasphaera elsdenii, Bacteroides monomorpha, Bacteroides acidogenes, and Bacteroides fragilis, in the treatment of glioblastoma, and the direct or indirect contributions of these microorganisms to the treatment of glioblastoma have not been fully explored. Therefore, studying the role of these probiotics in the progression and treatment of glioblastoma may provide a basis for the development of new therapeutic strategies.

[0006] Megasphaera elsdenii ( Megasphaera elsdenii) is a member of the Lactobacillus family and is an anaerobic Gram-negative bacterium that is primarily isolated from rumen animals such as cattle. It plays a vital role in the fermentation process in the rumen and is able to ferment a variety of carbohydrates such as lactose, fructose, and glucose. These metabolic activities are not only extremely important for the energy metabolism of animals, but also help regulate the pH of the rumen environment to prevent over-acidification, which is essential for the digestion and overall health of animals. Studies have shown that Megasphaera elsdenii is able to regulate lactic acid metabolism and maintain a balance in lactic acid levels in the environment.

[0007] Bacteroides monomorpha ( Bacteroides uniformis ) is a Gram-negative, anaerobic, rod-shaped bacterium belonging to the genus Bacteroides, which mainly inhabits the intestines of humans and animals. This bacterium can grow in an anaerobic environment and ferment a variety of carbohydrates to produce short-chain fatty acids (SCFAs) such as propionate and acetate. It can also break down polysaccharides and cellulose to help the host extract additional energy and nutrients. It also plays an important role in maintaining the balance of intestinal flora, immune system function, and intestinal barrier maintenance. At present, literature has reported the positive role of Bacteroides monotheca in improving colon inflammation and regulating depressive-like behavior in mice through the gut-brain axis.

[0008] Bacteroides acidogenes ( Bacteroides acidifaciens ) is an anaerobic bacterium belonging to the Bacteroidetes family, which is mainly found in the intestines of mammals. This bacterium is an important component of the intestinal microbiota and is involved in maintaining intestinal health and metabolic balance. It helps maintain the acid-base balance of the intestine by producing acidic substances such as short-chain fatty acids, providing energy for intestinal cells, while regulating the immune system and reducing inflammation. Studies have shown that in certain disease states, such as inflammatory bowel disease, the number of acid-producing Bacteroides may change, showing that it is closely related to the health of the intestine.

[0009] Bacteroides fragilis ( Bacteroides fragilis ) is an anaerobic, Gram-negative bacterium that is widely found in the intestines of humans and other warm-blooded animals. Although this bacterium only accounts for a small proportion of the intestinal microbiota, it has an important impact on intestinal health, mainly regulating the host immune system through its secreted polysaccharide A (PSA), promoting immune tolerance and anti-inflammatory state. Precisely because of its contribution to immune regulation, Bacteroides fragilis is being studied as a potential tool for the treatment of autoimmune and other inflammatory diseases.

[0010] However, there are no reports on the use of Megasphaera elsdenii, Bacteroides monomorpha, Bacteroides acidogenes, and Bacteroides fragilis in the treatment of glioblastoma. Summary of the invention

[0011] In order to solve the deficiencies of the prior art, the purpose of the present invention is to provide a pharmaceutical composition for treating glioblastoma, which mainly involves the use of four intestinal probiotics (Giant coccus elsdenii, Bacteroides monotheca, Bacteroides acidogenicus, and Bacteroides fragilis) in the preparation of drugs for treating glioblastoma. The present invention has proved through a large number of experiments that the pharmaceutical composition includes one or more of Giant coccus elsdenii, Bacteroides monotheca, Bacteroides acidogenicus, and Bacteroides fragilis, which can effectively play an auxiliary therapeutic role on glioblastoma.

[0012] In order to achieve the above object, the present invention adopts the following technical solution:

[0013] In a first aspect, the present invention provides four strains of intestinal probiotics having an adjuvant therapeutic effect on glioblastoma, wherein the Megasphaera elsdenii ( Megasphaera elsdenii ) is deposited as ATCC 25940, the Bacteroides monomorpha ( Bacteroides uniformis ) is deposited as ATCC 8492, the acidogenic Bacteroides ( Bacteroides acidifaciens ) is deposited as JCM 10556, the Bacteroides fragilis ( Bacteroides fragilis ) is deposited with ATCC 25285.

[0014] The present invention provides four intestinal probiotics with auxiliary therapeutic effects on glioblastoma, all of which have anti-tumor effects, significantly inhibit tumor growth in a mouse glioblastoma tumor model, and regulate intestinal microecology. Application of Giant Coccus elsdenii, Bacteroides monomorpha, Bacteroides acidogenes, and Bacteroides fragilis in the preparation of drugs for treating brain glioblastoma.

[0015] A pharmaceutical composition for treating glioblastoma, comprising one or more of Bacteroides monocytogenes, Bacteroides acidogenes, Bacteroides fragilis and Megasphaera elsdenii.

[0016] Furthermore, the pharmaceutical composition contains Megasphaera elsdenii.

[0017] Furthermore, the pharmaceutical composition contains Bacteroides monotheca, Bacteroides acidogenicus, and Bacteroides fragilis.

[0018] Furthermore, the pharmaceutical composition contains Megasphaera elsdenii, Bacteroides monomorpha, Bacteroides acidogenicus and Bacteroides fragilis.

[0019] Furthermore, the Bacteroides monomorpha, Bacteroides acidogenicus, Bacteroides fragilis, Megasphaera elsdenii are in the form of one or more of their derivatives, bacterial cell lysates or culture supernatants.

[0020] Furthermore, the derivative is a strain obtained by genetic recombination, transformation or modification, attenuation, chemical treatment, physical treatment or inactivation of the above strain.

[0021] The present invention also provides four intestinal probiotics for anti-tumor (Giant coccus elsdenii, Bacteroides monomorpha, Bacteroides acidogenicus, Bacteroides fragilis). The appropriate dosage of the bacterial flora is 10 6 -10 10 CFU / ml, preferably 10 9 CFU / ml.

[0022] Furthermore, when the pharmaceutical composition is composed of multiple bacteria, the number of cells of each bacteria is the same.

[0023] Furthermore, the glioblastoma includes: astrocytoma, oligodendroglioma, ependymoma, anaplastic astrocytoma, pleomorphic variant of glioblastoma, and papillary glioblastoma.

[0024] The beneficial effects achieved by one or more technical solutions of the present invention are:

[0025] Glioblastoma is a malignant brain tumor, and its treatment methods include surgical resection, radiotherapy, chemotherapy, and targeted therapy. However, due to the high recurrence rate and treatment side effects of glioblastoma, how to improve the quality of life of patients and prolong survival has become a research focus. Recent treatment advances include emerging strategies such as immunotherapy and cell therapy, which have improved the treatment effect to a certain extent, but still face many challenges. Glioblastoma patients often encounter many side effects during traditional chemotherapy and radiotherapy, such as nausea and diarrhea. These symptoms not only affect the treatment effect of patients, but also may reduce the patient's immune response rate to drugs due to intestinal flora imbalance. The four intestinal probiotics proposed in the present invention (Giant Coccus elsdenii, Bacteroides monomorpha, Bacteroides acidogenicus, Bacteroides fragilis) have important application potential in the adjuvant treatment of glioblastoma. The application of these four probiotics can not only improve the treatment effect of glioblastoma patients, but also improve the quality of life of patients by maintaining intestinal health. This new strategy provides a promising auxiliary means for the comprehensive treatment of glioblastoma and is expected to become an important part of the future treatment of glioblastoma. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] The accompanying drawings in the specification, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0027] Figure 1 shows the plate colony morphology of four intestinal probiotics, among which A is Giant coccus elsdenii, B is Bacteroides monomorpha, C is Bacteroides acidogenes, and D is Bacteroides fragilis.

[0028] Figure 2 Schematic diagram of the experiment to test the effects of four intestinal probiotics (Giantococcus elsdenii, Bacteroides monotheca, Bacteroides acidogenes, and Bacteroides fragilis) in the treatment of glioblastoma in a mouse glioblastoma model.

[0029] Figure 3 GBM#P3 luci Bioluminescence imaging of tumor-bearing mice. In vivo imaging images of small animals obtained by measuring the bioluminescence signals 3 weeks after treatment (4th week after tumor cell implantation). A: sham operation group; B: tumor-bearing + PBS group; C: tumor-bearing + antibiotic group; D: tumor-bearing + antibiotics + Megasphaera elsdenii group; E: tumor-bearing + antibiotics + Bacteroides monocytogenes + Bacteroides acidogenics + Bacteroides fragilis group; F: tumor-bearing + antibiotics + Megasphaera elsdenii + Bacteroides monocytogenes + Bacteroides acidogenics + Bacteroides fragilis group.

[0030] Figure 4 GBM#P3 luci Bioluminescence imaging of tumor-bearing mice 3 weeks after treatment (4th week after tumor cell implantation) obtained by measuring the bioluminescence signal of the live imaging light flux comparison chart; A sham operation group; B tumor-bearing + PBS group; C tumor-bearing + antibiotics group; D tumor-bearing + antibiotics + Megasphaera elsdenii group; E tumor-bearing + antibiotics + Bacteroides monotheca + Bacteroides acidogenics + Bacteroides fragilis group, F tumor-bearing + antibiotics + Megasphaera elsdenii + Bacteroides monotheca + Bacteroides acidogenics + Bacteroides fragilis group.

[0031] Figure 5 Kaplan-Meier analysis was used to evaluate the overall survival of tumor-bearing nude mice treated with four intestinal probiotics compared with the control group. The log-rank test was used to evaluate the statistical significance of the differences, A sham operation group; B tumor-bearing + PBS group; C tumor-bearing + antibiotic group; D tumor-bearing + antibiotic + giant coccus elsdenii group; E tumor-bearing + antibiotic + Bacteroides monotheca + Bacteroides acidogenic + Bacteroides fragilis group; F tumor-bearing + antibiotic + giant coccus elsdenii + Bacteroides monotheca + Bacteroides acidogenic + Bacteroides fragilis group. DETAILED DESCRIPTION

[0032] The technical scheme of the present invention will be further described in detail below in conjunction with specific embodiments. It should be understood that the following embodiments are only exemplary descriptions and explanations of the present invention, and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are included in the scope that the present invention is intended to protect.

[0033] Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0034] Unless defined otherwise or clearly indicated by the context, all technical and scientific terms in the present disclosure have the same meaning as commonly understood by one of ordinary skill in the art to which the present disclosure belongs.

[0035] The tumor cells GBM#P3 used in this application were donated by Professor Rolf Bjerkvig of the University of Bergen, Norway. They are primary glioblastoma cells. The cell line GBM#P3 stably expressing luciferase (abbreviated as luci) was constructed. luci (Luciferase lentivirus was purchased from Heyuan Biotechnology (Shanghai) Co., Ltd.), which has in vivo imaging properties and was used to construct a glioblastoma xenograft model and further monitor intracranial tumor growth using small animal imaging equipment.

[0036] Example 1 Cultivation of Megasphaera elsdenii

[0037] Cultivation methods

[0038] Step 1: Freeze-dried Megasphaera elsdenii ( Megasphaera elsdenii ) strain, Megasphaera elsdenii ( Megasphaera elsdenii ) is deposited as ATCC25940. The strain was resuspended in 200 μl of TSB (Tryptone Soy Broth) medium, and 20 μl was taken from it and inoculated on Columbia blood agar plates for streaking. The colony morphology of the plate is shown in Figure 1 A. Afterwards, the plate was placed in an anaerobic jar equipped with a precise gas control system and incubated anaerobically at 37 degrees Celsius for 48 hours.

[0039] Step 2: After the initial culture, select healthy monoclonal colonies and inoculate them into 10 ml TSB (Tryptone Soy Broth) medium. Place the inoculated medium in an anaerobic environment at 37 degrees Celsius and continue to culture for 12 hours.

[0040] Step 3: Prepare 500 ml of TSB medium and add 1% of the total volume of the bacteria obtained in step 2. Continue culturing at the same 37 degrees Celsius anaerobic condition for 48 hours.

[0041] Step 4: After the culture is completed, centrifuge at 4 degrees Celsius and 6000 rpm for 10 minutes to precipitate the bacteria.

[0042] Step 5: Wash the cells twice with sterile PBS to remove non-cell-attached impurities. Finally, resuspend the cells with sterile PBS for subsequent long-term storage and viable count analysis.

[0043] Example 2 Bacteroides monomorpha culture

[0044] Cultivation methods

[0045] Step 1: Freeze-dried Bacteroides monomorpha purchased from Beijing Bio-Bowei Biotechnology Co., Ltd. Bacteroides uniformis ) strains, Bacteroides monomorpha ( Bacteroides uniformis ) is deposited as ATCC8492. The strain was resuspended in 200 μl of TSB (Tryptone Soy Broth) medium, and 20 μl was taken from it and inoculated on TSA+5% defibrinated sheep blood agar plates for streaking. The colony morphology on the plate is shown in Figure 1 B. Afterwards, the plate was placed in an anaerobic jar equipped with a precise gas control system and incubated anaerobically at 37 degrees Celsius for 48 hours.

[0046] Step 2: After the initial culture, select healthy monoclonal colonies and inoculate them into 10 ml TSB (Tryptone Soy Broth) medium. Place the inoculated medium in an anaerobic environment at 37 degrees Celsius and continue to culture for 12 hours.

[0047] Step 3: Prepare 500 ml of TSB medium and add 1% of the total volume of the bacteria obtained in step 2. Continue culturing at the same 37 degrees Celsius anaerobic condition for 48 hours.

[0048] Step 4: After the culture is completed, centrifuge at 4 degrees Celsius and 6000 rpm for 10 minutes to precipitate the bacteria.

[0049] Step 5: Wash the cells twice with sterile PBS to remove non-cell-attached impurities. Finally, resuspend the cells with sterile PBS for subsequent long-term storage and viable count analysis.

[0050] Example 3 Cultivation of acidogenic Bacteroides

[0051] Cultivation methods

[0052] Step 1: Freeze-dried Bacteroides acidophilus (Bacteroides oxytoca) was purchased from Beijing Bio-Bowei Biotechnology Co., Ltd. Bacteroides acidifaciens ) strains, Bacteroides acidophilus ( Bacteroides acidifaciens ) is deposited as JCM 10556. The strain was resuspended in 200 μl of GAM medium, and 20 μl was taken from it and inoculated on a modified GAM agar plate for streaking. The colony morphology of the plate is shown in Figure 1 C. Afterwards, the plate was placed in an anaerobic jar equipped with a precise gas control system and incubated anaerobically at 37 degrees Celsius for 48 hours.

[0053] Step 2: After the initial culture, select healthy monoclonal colonies and inoculate them into 10 ml TSB (Tryptone Soy Broth) medium. Place the inoculated medium in an anaerobic environment at 37 degrees Celsius and continue to culture for 12 hours.

[0054] Step 3: Prepare 500 ml of TSB medium and add 1% of the total volume of the bacteria obtained in step 2. Continue culturing at the same 37 degrees Celsius anaerobic condition for 48 hours.

[0055] Step 4: After the culture is completed, centrifuge at 4 degrees Celsius and 6000 rpm for 10 minutes to precipitate the bacteria.

[0056] Step 5: Wash the cells twice with sterile PBS to remove non-cell-attached impurities. Finally, resuspend the cells with sterile PBS for subsequent long-term storage and viable count analysis.

[0057] Example 4 Bacteroides fragilis culture

[0058] Cultivation methods

[0059] Step 1: Freeze-dried Bacteroides fragilis ( Bacteroides fragilis ) strain, Bacteroides fragilis ( Bacteroides fragilis ) is deposited as ATCC25285. The strain was resuspended in 200 μl of GAM medium, and 20 μl was taken from it and inoculated on TSA+5% defibrinated sheep blood agar plates for streaking. The colony morphology of the plate is shown in Figure 1 D. Afterwards, the plate was placed in an anaerobic jar equipped with a precise gas control system and incubated anaerobically at 37 degrees Celsius for 48 hours.

[0060] Step 2: After the initial culture, select healthy monoclonal colonies and inoculate them into 10 ml TSB (Tryptone Soy Broth) medium. Place the inoculated medium in an anaerobic environment at 37 degrees Celsius and continue to culture for 12 hours.

[0061] Step 3: Prepare 500 ml of TSB medium and add 1% of the total volume of the bacteria obtained in step 2. Continue culturing at the same 37 degrees Celsius anaerobic condition for 48 hours.

[0062] Step 4: After the culture is completed, centrifuge at 4 degrees Celsius and 6000 rpm for 10 minutes to precipitate the bacteria.

[0063] Step 5: Wash the cells twice with sterile PBS to remove non-cell-attached impurities. Finally, resuspend the cells with sterile PBS for subsequent long-term storage and viable count analysis.

[0064] Example 5 Construction of GF (Germ-Free) Animal Model

[0065] Male Balb / C mice aged 5 weeks were selected, and the mice were all in a specific pathogen-free state. During the experiment, the mice received the following antibiotics by oral gavage every day: vancomycin 100 mg / kg, neomycin sulfate 200 mg / kg, metronidazole 200 mg / kg, and ampicillin 200 mg / kg, with a dose of 200 microliters per oral gavage per mouse, and this treatment lasted for one week. The purpose of this program is to eliminate the microbiota in the intestines of mice and adjust the composition and structure of the original intestinal microbiota in mice, so as to provide conditions for subsequent intestinal probiotic transplantation.

[0066] Example 6 Test method

[0067] Six groups were set up, with 3 mice in each group, namely:

[0068] Sham operation group: 5-week-old male Balb / C mice were anesthetized with chloral hydrate and fixed on a stereotaxic apparatus. After disinfection, the scalp was incised, with the bregma point as the zero point, 1 mm back, 2 mm to the right, and 1.5 mm deep. The scalp was sutured after completion.

[0069] Tumor-bearing + PBS group: 5-week-old male Balb / C mice were anesthetized with chloral hydrate and fixed on a stereotaxic apparatus. After disinfection, the scalp was incised, with the bregma point as the zero point, 1 mm back, 2 mm to the right, and 1.5 mm deep. 6 / 5 μL tumor cells were slowly injected into the right anterior bregma of the mice, and the scalp was sutured after completion. Seven days after the injection of tumor cells, 200 μL of sterile PBS was directly orally administered to the mice, and the tube feeding operation was performed once every two days for 3 consecutive weeks.

[0070] Tumor-bearing + antibiotic group: 5-week-old animal model mice constructed in Example 5 were taken and anesthetized with chloral hydrate. The mice were fixed in a brain stereotaxic instrument. After disinfection, the scalp was incised. The bregma point was taken as the zero point, and the scalp was incised 1 mm backward, 2 mm to the right, and 1.5 mm deep. 6 / 5μl tumor cells were slowly injected into the right anterior bregma of the mouse, and the scalp was sutured after completion.

[0071] Tumor-bearing + antibiotics + Megasphaera elsdenii group: see Figure 2 In addition to the operation of the tumor-bearing + antibiotic group, the following steps were added: 7 days after the injection of tumor cells, the bacterial suspension was prepared using the Megasphaera elsdenii cultured in Example 1, and the total concentration of the bacterial suspension was adjusted to 10 9 CFU / ml. The prepared bacterial suspension was directly orally administered to mice at a dose of 200 μl by gavage. The gavage operation was performed every two days for 3 consecutive weeks.

[0072] Tumor-bearing + antibiotics + Bacteroides acidophilus + Bacteroides fragilis + Bacteroides monomorpha group: see Figure 2In addition to the operation of the tumor-bearing + antibiotic group, the following steps were added: 7 days after the injection of tumor cells, the Bacteroides monomorpha, Bacteroides acidogenes, and Bacteroides fragilis cultured in Examples 2, 3, and 4 were used to prepare bacterial suspensions, and the above three bacterial suspensions were mixed in a ratio of 1:1:1 in terms of bacterial cell count, and the total concentration of the bacterial suspension was adjusted to 10 9 CFU / ml. The prepared bacterial suspension was then orally administered to mice at a dose of 200 microliters by gavage. The gavage operation was performed every two days for 3 consecutive weeks.

[0073] Tumor-bearing + antibiotics + Megasphaera elsdenii + Bacteroides acidophilus + Bacteroides fragilis + Bacteroides monomorpha group: see Figure 2 In addition to the operation of the tumor-bearing + antibiotic group, the following steps were added: 7 days after the injection of tumor cells, the bacterial suspension was prepared using the giant coccus of elsden, the monomorphic Bacteroides, the acidogenic Bacteroides, and the fragilis cultured in Examples 1, 2, 3, and 4, and the above four bacterial suspensions were mixed in a ratio of 1:1:1:1 in terms of bacterial cell count, and the total concentration of the bacterial suspension was adjusted to 10 9 CFU / ml. The prepared bacterial suspension was then orally administered to mice at a dose of 200 microliters by gavage. The gavage operation was performed every two days for 3 consecutive weeks.

[0074] After the animal model was cultured according to the above protocol, the bioluminescence value of the tumor was quantified using the Living Image 2.5 software package (Xenogen) and subsequent statistics were performed.

[0075] Results Analysis

[0076] in, Figure 3 GBM#P3 luci Bioluminescent imaging of tumor-bearing mice. The bioluminescent signal was measured in the small animal live imaging image 3 weeks after treatment (4 weeks after tumor cell implantation). The coordinate axis on the right shows the intensity of the bioluminescent signal, and its scale and value reflect the changes in different luminescent intensity ranges. It can be seen from the small animal live imaging image that GBM#P3 luci Strong cell proliferation ability, GBM#P3 luci Four weeks after tumor cell implantation, compared with the sham operation group (A), the bioluminescent signal intensity of the tumor-bearing + PBS group (B), tumor-bearing + antibiotic group (C), tumor-bearing + antibiotic + Megasphaera elsdenii group (D), tumor-bearing + antibiotic + Bacteroides fragilis + Bacteroides monotheca group (E), and tumor-bearing + antibiotic + Megasphaera elsdenii + Bacteroides fragilis + Bacteroides monotheca group (F) showed a significant increase trend, and the overall bioluminescent signal intensity remained at 10 6 Up to 10 8 (p / sec / cm 2The bioluminescence signal intensity of the tumor-bearing + PBS group (B) and the tumor-bearing + antibiotic group (C) was maintained at an average level of 10 8 The bioluminescent signal intensity of the tumor-bearing + antibiotics + Megasphaera elsdenii group (D), tumor-bearing + antibiotics + Bacteroides fragilis + Bacteroides monomorpha group (E), and tumor-bearing + antibiotics + Megasphaera elsdenii + Bacteroides fragilis + Bacteroides monomorpha group (F) was maintained at an average of 10 7 Further analysis of the bioluminescence intensity of each tumor-forming group showed that by the fourth week after cell implantation, compared with the two tumor-bearing control groups (tumor-bearing + PBS group (B), tumor-bearing + antibiotic group (C)), the three treatment groups (tumor-bearing + antibiotics + Megasphaera elsdenii group (D), tumor-bearing + antibiotics + acidogenic Bacteroides + Bacteroides fragilis + Bacteroides monotheca group (E), and tumor-bearing + antibiotics + Megasphaera elsdenii + acidogenic Bacteroides + Bacteroides fragilis + Bacteroides monotheca group (F)) showed significant therapeutic effects, specifically, the tumor size of the treatment group was significantly smaller than that of the two control groups.

[0077] in, Figure 4 GBM#P3 luci Bioluminescent imaging of tumor-bearing mice. Comparison of in vivo imaging fluorescence flux obtained by measuring bioluminescent signals 3 weeks after treatment (4th week after tumor cell implantation). From the fluorescence flux comparison chart, it can be seen that in the 4th week after cell implantation, compared with the two tumor-bearing control groups (tumor-bearing + PBS group (B), tumor-bearing + antibiotic group (C)), the tumor size of the three treatment groups (tumor-bearing + antibiotics + Megasphaera elsdenii group (D), tumor-bearing + antibiotics + acidogenic Bacteroides + Bacteroides fragilis + Bacteroides monotheca group (E) and tumor-bearing + antibiotics + Megasphaera elsdenii + acidogenic Bacteroides + Bacteroides fragilis + Bacteroides monotheca group (F)) was significantly smaller than that of the two control groups, as shown by the fluorescence signal intensity of the treatment group being about 100 times smaller than that of the control group.

[0078] in, Figure 5 Kaplan-Meier analysis was performed to evaluate the overall survival of tumor-bearing nude mice treated with the four intestinal probiotics compared with the control group. The log-rank test was used to assess the statistical significance of the differences. luci By bioluminescence imaging of the survival time of tumor-bearing mice, we found that compared with the two tumor-bearing control groups (tumor-bearing + PBS group (B), tumor-bearing + antibiotics group (C)), the survival time of tumor-bearing mice in the three treatment groups (tumor-bearing + antibiotics + Megasphaera elsdenii group (D), tumor-bearing + antibiotics + Bacteroides fragilis + Bacteroides monotheca group (E), and tumor-bearing + antibiotics + Megasphaera elsdenii + Bacteroides fragilis + Bacteroides monotheca group (F)) was significantly prolonged.

[0079] Notably, tumor-bearing mice treated with M. elsdenii showed a better survival percentage than those treated with B. acidophilus, B. fragilis, and B. monotheca, suggesting that M. elsdenii has specific bioactive components or biological mechanisms that interact with tumor cells, leading to its more effective antitumor effect.

[0080] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A pharmaceutical composition for treating glioblastoma, characterized in that: The pharmaceutical composition consists of Bacteroides monocytogenes, Bacteroides acidogens, Bacteroides fragilis and Megasphaera elsdenii, wherein the Megasphaera elsdenii is deposited with ATCC25940, the Bacteroides monocytogenes is deposited with ATCC8492, the Bacteroides acidogens is deposited with JCM10556, and the Bacteroides fragilis is deposited with ATCC25285.

2. A pharmaceutical composition for treating glioblastoma according to claim 1, characterized in that: The concentration of the bacterial flora in the pharmaceutical composition is 10 6 -10 10 CFU / mL.

3. A pharmaceutical composition for treating glioblastoma according to claim 2, characterized in that: The concentration of the bacterial flora in the pharmaceutical composition is 10 9 CFU / mL.

4. A pharmaceutical composition for treating glioblastoma according to claim 1, characterized in that: The number of cells of each bacterium in the pharmaceutical composition is the same.

5. Use of a pharmaceutical composition in the preparation of a drug for treating glioblastoma, wherein the pharmaceutical composition is composed of Bacteroides monocytogenes, Bacteroides acidophilus, Bacteroides fragilis and Megasphaera elsdenii, wherein the Megasphaera elsdenii has a preservation number of ATCC25940, the Bacteroides monocytogenes has a preservation number of ATCC 8492, the Bacteroides acidophilus has a preservation number of JCM10556, and the Bacteroides fragilis has a preservation number of ATCC 25285.

6. The use according to claim 5, characterized in that The concentration of the bacterial flora in the pharmaceutical composition is 10 6 -10 10 CFU / mL.

7. The use according to claim 6, characterized in that The concentration of the bacterial flora in the pharmaceutical composition is 10 9 CFU / mL.

8. The use according to claim 5, characterized in that: The number of cells of each bacterium in the pharmaceutical composition is the same.

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