Ruminococcus torsionosus with effect of inhibiting colorectal cancer and application of ruminococcus torsionosus

Abstract

The invention discloses ruminococcus torsionosus with a colorectal cancer inhibiting effect and application thereof, creatively and accidentally discovers that the ruminococcus torsionosus has the colorectal cancer inhibiting effect for the first time, and lays a key experimental foundation for subsequent development of a colorectal cancer micro-ecological treatment strategy taking the ruminococcus torsionosus as a core. The important theoretical innovation significance and clinical transformation potential are realized.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biomedicine, and particularly relates to a Ruminococcus torques with an inhibiting effect on colorectal cancer and an application thereof. BACKGROUND

[0002] Colorectal cancer (CRC) is one of the malignant tumors with high morbidity and mortality worldwide, and its occurrence and development are closely related to intestinal flora. Intestinal flora imbalance not only damages the integrity of intestinal mucosal barrier and triggers chronic inflammatory response, but also regulates host cell proliferation and apoptosis through metabolic products, thereby affecting the occurrence process of tumor. Although existing studies show that certain intestinal microorganisms may be related to colorectal cancer, the role of Ruminococcus torques in colorectal cancer is still unclear, and existing literature focuses on the relationship between other flora (such as Fusobacterium nucleatum and Bifidobacterium) and colorectal cancer. There is no clear conclusion on whether Ruminococcus torques participates in the pathological process of colorectal cancer and what kind of regulatory role it plays, and no study has confirmed that it can directly inhibit the growth of colorectal cancer.

[0003] In addition, numerous existing technologies give the reverse technical teaching that "Ruminococcus torques may be able to induce the occurrence of colorectal cancer". Specifically, the existing technology (Wu Y, Zhuang J, Zhang Q, et al. Aging characteristics of colorectal cancer based on gut microbiota [J]. Cancer Medicine, 2023, 12(17): 17822-17834.) explicitly discloses that Ruminococcus torques is enriched in colorectal cancer; the existing technology (Valentino V, De Filippis F, Marotta R, et al. Genomic features and prevalence of Ruminococcus species in humans are associated with age, lifestyle, and disease [J]. Cell Reports, 2024, 43(12).) also explicitly discloses that high levels of Ruminococcus torques are related to the occurrence and development of colorectal cancer. Ruminococcus torques can destroy the integrity of the intestinal barrier by degrading mucin, thereby promoting the invasion of pathogens and triggering inflammation and disease. It can be seen that the above existing technologies explicitly give the reverse technical teaching that "Ruminococcus torques may be able to induce the occurrence of colorectal cancer, rather than treat colorectal cancer".

[0004] However, the present application first creatively and unexpectedly discovers that the Ruminococcus torques has an inhibitory effect on colorectal cancer through the dual verification of in vitro experiments and in vivo experiments, which not only provides the first direct evidence for the role of the Ruminococcus torques in the occurrence and development of colorectal cancer, but also breaks through the existing research boundaries of intestinal flora intervention in colorectal cancer, lays a key experimental foundation for the subsequent development of a colorectal cancer microecological treatment strategy (such as probiotic preparations, flora transplantation programs, etc.) with the Ruminococcus torques as the core, and has important theoretical innovation significance and clinical transformation potential. SUMMARY

[0005] In order to make up for the above-mentioned technical research blank in the prior art, the purpose of the present application is to provide a Ruminococcus torques having an inhibitory effect on colorectal cancer and applications thereof.

[0006] The present application achieves the above-mentioned application purposes by adopting the following technical solutions:

[0007] The first aspect of the present application provides an application of the Ruminococcus torques in the preparation of a drug for treating and / or preventing colorectal cancer.

[0008] Further, the sequence of the Ruminococcus torques is shown in SEQ ID NO. 1.

[0009] Further, the Ruminococcus torques can inhibit the proliferation of colorectal cancer cells and / or promote the apoptosis of colorectal cancer cells.

[0010] Further, the drug further comprises a pharmaceutically acceptable carrier;

[0011] Alternatively, the pharmaceutically acceptable carrier is a filler, an enteric coating material, an adhesion agent, a stabilizer, a diluent, a suspending agent, a bacteriostatic agent, a rectal preparation carrier, an intratumoral implant preparation carrier, a prebiotic carrier or an inert carrier.

[0012] In some embodiments, the filler, also known as a diluent, is used to increase the volume of the preparation, improve the formability, and does not affect the activity of the strain, for example, lactose, sucrose: natural sugars, high safety, can provide a small amount of nutrients for the strain, suitable for preparing ordinary live bacteria capsules, powders; malt dextrin, soluble starch: easy to dissolve, non-irritating, can be stably mixed with the strain, suitable for freeze-dried oral powders, compound preparations (used with prebiotics); microcrystalline cellulose: good fluidity and formability, can be used for the preparation of enteric-coated tablets and enteric mucosa-adhesive tablets to avoid inactivation of the strain during preparation.

[0013] In some embodiments, the enteric coating material is used to prepare enteric formulations (enteric capsules, enteric pellets) to protect the bacterial strain from the strong acid environment in the stomach, for example, acrylic resins (Eudragit L100, Eudragit S100): insoluble in the stomach (pH < 3), soluble in the small intestine (pH > 5.5) or colon (pH > 7), achieving targeted release; cellulose acetate phthalate (CAP): a traditional enteric material, good film-forming property, can be used to coat tablets or pellets, ensuring that the live strain enters the intestine; shellac: a natural enteric material, high safety, suitable for preparing prophylactic live bacterial preparations that need to be taken for a long time.

[0014] In some embodiments, the adhesion agent, also known as mucosal adhesion material, is used in intestinal mucosal adhesion formulations (such as adhesion microspheres, adhesion tablets) to prolong the contact time of the bacterial strain with the intestinal mucosa and promote colonization, for example, chitosan: a natural cationic polysaccharide that can bind to the intestinal mucosa (negatively charged) and has certain probiotic effects, which can assist in improving the intestinal microecology; hydroxypropyl methylcellulose (HPMC): moderate adhesion, can form a gel layer to make the preparation adhere to the surface of the intestinal mucosa and slowly release the bacterial strain; carbomer: good mucosal adhesion and swelling, suitable for preparing colon-targeted adhesion formulations to increase the concentration of the bacterial strain in the lesion site of the colon.

[0015] In some embodiments, the stabilizer, also known as a protective agent, is directed against the live characteristics of the T. praefermentans strain to prevent the strain from being inactivated during storage and transportation, especially for freeze-dried preparations (freeze-dried bacterial powder, freeze-dried capsules), for example, freeze-drying protective agents: sucrose, trehalose, mannitol, which can maintain the cell structure of the bacterial strain during freeze-drying and reduce ice crystal damage; antioxidants: vitamin C, sodium sulfite, which can prevent the strain from being inactivated due to oxidative stress, suitable for compound preparations containing the bacterial strain (combined with chemotherapy drugs to reduce the oxidative damage of the drugs to the bacterial strain); buffers: phosphate buffer (PBS), citrate buffer, which can adjust the pH of the preparation to be close to the intestinal environment (pH 6.5-7.5) to maintain the activity of the bacterial strain.

[0016] In some embodiments, the diluent, also known as a solvent, is used to dissolve or suspend the bacterial strain to ensure the flowability and safety during injection, for example, sterile normal saline (0.9% sodium chloride injection): which can be directly used as a carrier for bacterial injection suspension without affecting the ability of the bacterial strain to enter tumor cells; sterile water for injection: which is used to reconstitute freeze-dried injection powders, and needs to be combined with a buffer to adjust the pH to avoid inactivation of the bacterial strain during reconstitution; phosphate buffered saline (PBS, pH 7.2-7.4): which has a pH close to that of human body fluids, good biocompatibility, can reduce local irritation during intratumoral injection, and is suitable for long-term multiple injections.

[0017] In some embodiments, the suspending agent, also known as suspending agent, if the strain exists in the form of particles (such as microencapsulated strains), is used to maintain the suspended state, avoid sedimentation, and ensure uniform injection dose. For example, polysorbate 80 (Tween 80): a non-ionic surfactant that can reduce interfacial tension, uniformly disperse strain particles, and has high safety and no obvious toxic side effects; sodium carboxymethyl cellulose (CMC-Na): moderate viscosity that can increase the viscosity of the injection to prevent strain particles from settling, and adapt to the local retention requirements of intratumoral injection; sodium hyaluronate: a natural polysaccharide with good biocompatibility and biodegradability, which can be used as a suspending agent for intratumoral injection preparations, and can form a gel in the tumor microenvironment to prolong the action time of the strain.

[0018] In some embodiments, the bacteriostatic agent is only used for multi-dose injections. If a multi-dose intratumoral injection preparation is prepared, a bacteriostatic agent needs to be added to prevent microbial contamination without affecting the activity of the strain. For example, benzyl alcohol: a commonly used injection bacteriostatic agent with a concentration of 0.5%-1% that has no obvious inhibitory effect on the strain and can reduce local pain during injection; chlorobutanol: has bacteriostatic and local anesthetic effects, and is suitable for the reconstitution carrier of multi-dose lyophilized injection powders to ensure safety for multiple uses.

[0019] In some embodiments, the rectal preparation (suppository / gel) carrier adapts to the scenario of rectal local tumor treatment, and needs to melt or dissolve in the rectum to release the strain and contact the lesion. For example, suppository base: cocoa butter, semi-synthetic fatty acid glycerides (such as coconut oil ester), which are solid at room temperature, rapidly melt in the rectum (about 37°C), release the strain, and have no irritation; gel base: carbomer 940, hydroxypropyl methylcellulose (HPMC), which can form a semi-solid gel, adhere to the rectal mucosa surface, prolong the action time of the strain, and is suitable for postoperative adjuvant therapy of rectal cancer.

[0020] In some embodiments, the intratumoral implant preparation (microspheres, implant tablets) carrier needs to be biodegradable and slowly release the strain. For example, degradable polymers: poly(lactic-co-glycolic acid) (PLGA), polylactic acid (PLA), which can be made into microspheres or implant tablets, slowly degrade after implantation into the tumor (degradation period 1-3 months), continuously release the strain, and reduce repeated injections; natural high molecular materials: sodium alginate, gelatin, which have good biocompatibility and can be prepared into microspheres by ionic crosslinking method, implanted into the tumor after loading the strain, and have no toxicity of degradation products, and are suitable for local control of advanced colorectal cancer.

[0021] In some embodiments, the prebiotic carrier includes but is not limited to: fructooligosaccharides, galactooligosaccharides, which can not only be used as fillers for preparations, but also can promote the proliferation of strains in the intestinal tract and enhance their inhibitory effect on colorectal cancer.

[0022] In some embodiments, the inert carrier includes, but is not limited to, talc, magnesium stearate, a lubricant for tablets or capsules, reducing frictional damage to the strain during the preparation process and equipment, and ensuring the survival rate of the strain.

[0023] Further, the dosage form of the drug is an oral preparation, an injection preparation, an enteric preparation, a colon-targeted preparation, a rectal preparation, an intratumoral implant preparation, a freeze-dried preparation, a viable capsule preparation, or a microcapsule preparation.

[0024] In some embodiments, the oral preparation uses Anaerotruncus colihominis with a sequence as shown in SEQ ID NO. 1 as the active ingredient, and can maintain the activity of the strain through a pharmaceutically acceptable carrier such as a filler (e.g., lactose, malt dextrin), a stabilizer (e.g., sucrose, trehalose), and the like. The administration method is convenient, can enter the intestinal tract through the digestive tract, plays a role in inhibiting colorectal cancer by regulating the intestinal microecology and repairing the intestinal mucosal barrier, can be adapted to the scenarios of colorectal cancer prevention and adjuvant therapy of intestinal tumors, and part of the oral preparations can be compounded with prebiotics to further promote the colonization of the strain in the intestine and enhance the efficacy.

[0025] In some embodiments, the injection preparation uses Anaerotruncus colihominis with a sequence as shown in SEQ ID NO. 1 as the core, is mainly administered through intratumoral injection, and commonly uses sterile normal saline, phosphate buffer (pH 7.2-7.4) as the solvent. If the strain is in a granular form, polysorbate 80 and the like can be added as a suspending agent to ensure uniform dispersion. This dosage form can directly deliver the strain to the inside of the tumor, allowing it to quickly enter tumor cells and regulate the expression of proliferation and apoptosis-related genes. Intratumoral injection of this preparation can significantly reduce the volume and weight of subcutaneous tumors in mice, and the selection of carriers follows the principle of non-toxicity and non-damage to the activity of the strain, which meets the safety and efficacy requirements of local anti-tumor treatment in the clinic.

[0026] In some embodiments, the enteric preparation includes enteric tablets, enteric capsules, and enteric pellets, and uses acrylic resin, cellulose acetate phthalate, and the like as the key enteric coating material to avoid the inactivation of Anaerotruncus colihominis in the strong acid environment in the stomach. It can ensure that the strain is alive and enters the small intestine and colon, and precisely acts on the colorectal tumor lesions in the intestinal tract or participates in the regulation of the intestinal microecology, especially for patients who are sensitive to the gastric environment and need to ensure the colonization rate of the strain, and provides a stable drug delivery route for intestinal local intervention for colorectal cancer.

[0027] In some embodiments, the colon-targeted preparation includes colon-targeted slow-release particles, colon-targeted gel microspheres, and triggers drug release using intracolonic specific enzymes (such as azo reductase) or pH gradient (pH 6.5-7.5) with microcrystalline cellulose, chitosan, etc. as carrier materials. This dosage form can precisely release the Streptococcus ruminantium in the colon, reduce the loss of the strain in the upper part of the intestinal tract, directly act on the colorectal tumor lesion, enhance the inhibitory effect on inflammation-induced colorectal cancer (such as AOM / DSS-induced model), effectively increase the concentration of the strain in the colon lesion, and adapt to the needs of targeted therapy for colorectal cancer (especially colon cancer).

[0028] In some embodiments, the rectal preparation includes rectal suppositories, rectal gels, and rectal creams, with cocoa butter, semi-synthetic fatty acid glycerides as the matrix of the suppositories, and carbomer, hydroxypropyl methyl cellulose as the matrix of the gels / creams. Through rectal administration, the Streptococcus ruminantium directly contacts the rectal tumor tissue or rectal mucosa, avoiding the degradation by digestive enzymes and the loss of blood circulation during oral administration, and is suitable for patients with tumors located in the lower rectum or postoperative intestinal function not recovered, who cannot take orally. It can play a targeted role in local anti-tumor effect in the rectum, and meet the needs of local treatment of rectal cancer.

[0029] In some embodiments, the intratumoral implant preparation includes degradable intratumoral implant microspheres and implant tablets, with biodegradable materials such as polylactic acid-glycolic acid copolymer (PLGA) and sodium alginate as carriers. After implanting the preparation loaded with Streptococcus ruminantium into the tumor, the carrier can be slowly degraded (degradation period 1-3 months) to continuously release the strain, so that the strain can play a role in the tumor microenvironment for a long time, reducing the operation of repeated intratumoral injection, and can inhibit tumor growth for a long time, which is suitable for local lesion control of advanced colorectal cancer.

[0030] In some embodiments, the freeze-dried preparation includes freeze-dried bacterial powder, freeze-dried oral powder, and freeze-dried injection powder, with sucrose, sodium alginate, mannitol, etc. as freeze-drying protectants. The freeze-drying process significantly improves the stability of Streptococcus ruminantium, prolongs the storage time (can be stored for 1-2 years at low temperature), and adapts to the needs of large-scale production and multiple clinical administration scenarios. Moreover, it can guarantee the activity of the strain during storage and transportation, and meets the requirements of convenience and stability of clinical medication.

[0031] In some embodiments, the live bacteria capsule preparation takes the sequence shown in SEQ ID NO. 1 as the active ingredient, and lactose and soluble starch as the fillers, with the capsule shell as the carrier material. The preparation process is relatively simple and low in cost, and is suitable for patients with upper intestinal tumors or mild intestinal microecological imbalance. If the patient takes an acid-suppressing drug, the pH value in the stomach is relatively high, which can ensure the survival of the strain into the intestine, and the strain can colonize in the intestine to inhibit colorectal cancer. The preparation can be used as a daily microecological preparation for the prevention of colorectal cancer.

[0032] In some embodiments, the microcapsule preparation encapsulates the Anaerotruncus colihominis in sodium alginate-chitosan and other microcapsule materials, and the particle size of the microcapsule can be controlled (1-10 μm) to protect the strain from gastric acid and digestive enzymes, and to control the release rate of the strain to achieve sustained release. When taken orally, the microcapsule can evenly distribute the strain in the intestine to improve the colonization efficiency. If the particle size meets the requirements of the vascular access, the microcapsule can also be injected intravenously to reach the metastatic foci of tumors throughout the body, thereby expanding the scope of action. The dosage form can enhance the stability and efficacy of the strain through the protection and controlled release of the carrier material, and meet the needs of multi-site intervention for primary and metastatic foci of colorectal cancer.

[0033] The second aspect of the present application provides an Anaerotruncus colihominis with an inhibitory effect on colorectal cancer.

[0034] Further, the sequence of the Anaerotruncus colihominis is shown in SEQ ID NO. 1.

[0035] The third aspect of the present application provides a pharmaceutical composition for treating and / or preventing colorectal cancer.

[0036] Further, the pharmaceutical composition comprises the Anaerotruncus colihominis according to the second aspect of the present application.

[0037] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

[0038] In some embodiments, the selection of the pharmaceutically acceptable carrier needs to strictly match the characteristics of the Anaerotruncus colihominis, i.e., “intestinal normal flora member, need to maintain the activity of live bacteria, and adapt to multiple scenarios of administration”, while meeting the safety, stability and functionality requirements.

[0039] In some embodiments, for oral pharmaceutical compositions (such as ordinary live bacteria capsules and enteric-coated tablets), lactose and maltodextrin can be used as fillers to adjust the volume and formability of the preparation, and acrylic acid resin and cellulose acetate phthalate can be used as enteric-coated materials to protect the strain from strong acid in the stomach and ensure that the strain colonizes in the intestine in the form of live bacteria.

[0040] In some embodiments, for injection-type pharmaceutical compositions (such as intratumoral injection preparations), sterile normal saline, phosphate buffer (pH 7.2-7.4) is used as the solvent to avoid stimulating the tumor microenvironment. If the strain is in granular form, a suspending agent such as polysorbate 80 can be added to ensure uniform injection dose.

[0041] In some embodiments, for rectal, intratumoral implantation, and other local administration compositions, cocoa butter (suppository base), polylactic acid-glycolic acid copolymer (PLGA, implant carrier), and the like can be used to allow the strain to directly act on the local tumor lesion and reduce transport loss.

[0042] In addition, in some embodiments, cryoprotectants such as sucrose, trehalose, and the like (suitable for freeze-dried preparations), prebiotic carriers such as fructooligosaccharides, and the like (promote strain proliferation) can be introduced. These carriers not only have no significant toxic side effects, but also can protect the activity of the strain, optimize the administration efficiency, enhance the colonization ability of the strain, further improve the therapeutic and prophylactic effects of the pharmaceutical composition on colorectal cancer, expand the dosage form range of the composition (such as oral, injection, local administration, etc.), and meet the clinical medication needs of different patients (such as postoperative patients who cannot take orally, rectal tumor patients, and advanced local lesion control patients).

[0043] The fourth aspect of the present application provides a pharmaceutical preparation for treating and / or preventing colorectal cancer.

[0044] Further, the pharmaceutical preparation comprises the pharmaceutical composition of the third aspect of the present application;

[0045] Optionally, the dosage form of the pharmaceutical preparation is an oral preparation, an injection preparation, an enteric preparation, a colon-targeted preparation, a rectal preparation, an intratumoral implantation preparation, a freeze-dried preparation, a viable bacteria capsule preparation, or a microcapsule preparation.

[0046] The fifth aspect of the present application provides any one of the following products:

[0047] (1) a bacterial agent for treating and / or preventing colorectal cancer, the bacterial agent comprising the Ruminococcus tortuosus of the second aspect of the present application;

[0048] (2) a food for treating and / or preventing colorectal cancer, the food comprising the Ruminococcus tortuosus of the second aspect of the present application;

[0049] (3) a health product for treating and / or preventing colorectal cancer, the health product comprising the Ruminococcus tortuosus of the second aspect of the present application;

[0050] (4) a dietary supplement for treating and / or preventing colorectal cancer, the dietary supplement comprising the Ruminococcus tortuosus of the second aspect of the present application;

[0051] (5) A diagnostic product for diagnosing colorectal cancer, comprising a reagent for detecting the abundance of the Ruminococcus torques of the second aspect of the present application.

[0052] In some embodiments, the probiotic is a microbial preparation that takes the specific Ruminococcus torques (the sequence is shown in SEQ ID NO. 1) of the second aspect of the present application as the core functional active ingredient, and is designed specifically for the treatment and prevention needs of colorectal cancer.

[0053] In some embodiments, the diagnostic product includes a detection kit, a detection test strip or a detection chip. Specifically, it mainly includes detection reagents such as a kit for extracting microbial DNA from samples (feces, intestinal tissue samples), specific primers and probes designed for Ruminococcus torques sequences (SEQ ID NO. 1), PCR reaction reagents (such as DNA polymerase, buffer), etc. Part of the product can also be matched with the operating instructions of the detection instrument (such as real-time fluorescent quantitative PCR instrument) to ensure the standardization of the detection process.

[0054] In some embodiments, the diagnostic product detects the relative abundance of Ruminococcus torques in the sample of the subject to be detected, and compares it with the standard abundance range of healthy people and colorectal cancer patients. If the abundance of the strain in the sample is significantly lower than the standard value of healthy people and close to or lower than the standard value of colorectal cancer patients, it indicates that there may be a risk of colorectal cancer, which needs to be further diagnosed by colonoscopy and pathological examination.

[0055] The sixth aspect of the present application provides a method for inhibiting the proliferation of colorectal cancer cells and / or promoting the apoptosis of colorectal cancer cells in vitro for non-therapeutic purposes, which comprises contacting a cell system in need with the Ruminococcus torques of the second aspect of the present application.

[0056] In addition, the present application also provides a method for diagnosing and / or assisting in diagnosing colorectal cancer, which comprises detecting the abundance of the Ruminococcus torques of the second aspect of the present application in a sample to be detected from a subject, and judging whether the subject is a colorectal cancer patient or has a risk of colorectal cancer based on the detection result.

[0057] In addition, the present application also provides a method for treating and / or preventing colorectal cancer, which comprises administering a therapeutically and / or preventively effective amount of the Ruminococcus torques of the second aspect of the present application, the pharmaceutical composition of the third aspect of the present application, the pharmaceutical preparation of the fourth aspect of the present application and / or the probiotic of the fifth aspect of the present application to a subject in need.

[0058] In some embodiments, the treatment and / or prevention refers to slowing, interrupting, arresting, alleviating, stopping, reducing, or reversing the progression or severity of an existing symptom, disorder, condition, or disease (e.g., colorectal cancer). Desirable effects include, but are not limited to, preventing occurrence or recurrence of disease, alleviating symptoms, diminishment of any direct or indirect pathological consequences of the disease, decreasing the rate of disease progression, ameliorating or palliating the disease state, and remission or improved prognosis.

[0059] In some embodiments, the route of administration of the pharmaceutical composition or pharmaceutical preparation of the present application is not limited as long as it can exert the desired therapeutic or prophylactic effect, including, but not limited to, topical, oral, cutaneous, intravenous, intra-arterial, intramuscular, intratracheal, subcutaneous, inhalation, enteral, rectal, etc. In some cases, it can be administered systemically, and in some cases, it can be administered locally.

[0060] In some embodiments, the dosage of the pharmaceutical composition or pharmaceutical preparation of the present application is appropriately prescribed according to the method of formulation, the mode of administration, the age, body weight, sex, state of illness, diet, administration time, route of administration, excretion rate, and reaction sensitivity of the patient, and the like, and generally, a skilled medical doctor can easily determine the prescription and the dosage of the administration effective for the desired treatment or prevention.

[0061] In some embodiments, the subject includes both human and non-human animals. Among them, the non-human animals include all vertebrates (e.g., mammals and non-mammals), such as non-human primates (e.g., cynomolgus monkeys), sheep, dogs, cows, chickens, amphibians, and reptiles. In some embodiments, the subject is preferably a human.

[0062] In some embodiments, the effective amount refers to an amount of the S. ruminantium of the second aspect of the present application, the pharmaceutical composition of the third aspect of the present application, the pharmaceutical preparation of the fourth aspect of the present application, and / or the bacterial agent of the fifth aspect of the present application sufficient to treat the specified disorder, condition, or disease (e.g., to ameliorate, alleviate, attenuate, and / or delay one or more of its symptoms).

[0063] In some embodiments, the effective amount is an amount sufficient to delay the development of a disease (e.g., colorectal cancer). In some embodiments, the effective amount is an amount sufficient to prevent or delay the recurrence of a disease. The effective amount can be administered in one or more administrations. The therapeutically effective amount of a pharmaceutical is dependent on various factors, including, but not limited to, the characteristics of the subject (e.g., height, weight, sex, age, and medication history), the severity of the disease, and the like.

[0064] The seventh aspect of the present application provides any of the following aspects:

[0065] (1) The use of the Ruminococcus torques in the preparation of a microbial agent, food, health product or dietary supplement for treating and / or preventing colorectal cancer according to the second aspect of the present application;

[0066] (2) The use of the reagent for detecting the abundance of the Ruminococcus torques according to the second aspect of the present application in the preparation of a diagnostic product for diagnosing colorectal cancer.

[0067] Compared with the prior art, the present application has the following advantages and beneficial effects:

[0068] (1) The present application first discovers that the Ruminococcus torques has an inhibitory effect on colorectal cancer. Specifically, the abundance of the Ruminococcus torques in the normal intestinal epithelial tissue of a colorectal cancer patient is significantly higher than that in the tumor tissue, and it is verified through animal experiments that the Ruminococcus torques can enter tumor cells, significantly inhibit the growth of subcutaneous tumors and the formation of spontaneous colorectal cancer induced by inflammation in mice, and reduce the number and volume of tumors. The new use of the Ruminococcus torques discovered for the first time provides a new microbial resource for the prevention and treatment of colorectal cancer, and has good application prospects;

[0069] (2) Strong targeting: The present application discovers through experiments that the Ruminococcus torques can enter tumor cells to play a role, directly affect the tumor microenvironment, and specifically inhibit the progression of colorectal cancer. After the strain enters the tumor cells, it can regulate related signaling pathways in the tumor cells, such as affecting the expression of cell proliferation and apoptosis-related genes, thereby inhibiting the growth of tumor cells, and has strong targeting;

[0070] (3) Natural safety: The Ruminococcus torques described in the present application belongs to a member of the normal intestinal flora, is isolated from healthy tissue, has no significant toxic and side effects, and is suitable for long-term application. After multiple generations of animal experiments, no adverse effects on the growth and development, physiological functions, etc. of the animals were found;

[0071] (4) High potential value: The present application provides a new target for microbial intervention of colorectal cancer, which can be developed into a new prevention or treatment preparation to make up for the shortcomings of existing chemical drugs. It can be used in combination with existing treatment methods to enhance the treatment effect, such as combined with chemotherapy drugs, which can reduce the toxic and side effects of the chemotherapy drugs, and at the same time improve the killing effect on tumor cells. BRIEF DESCRIPTION OF DRAWINGS

[0072] Figure 1 Distribution of Ruminococcus torques in normal intestinal epithelium and tumor tissue;

[0073] Figure 2 Effect of intratumoral injection of Ruminococcus torques on the growth of subcutaneous tumors in mice;

[0074] Figure 3: The effect of Ruminococcus torques on the tumorigenesis of AOM / DSS model mice by gavage;

[0075] Figure 4 : Cryo-EM image of MC38 cells co-cultured with Ruminococcus torques. DETAILED DESCRIPTION

[0076] The present application will be further described below in conjunction with specific examples. The following specific examples are only used to explain the present application and should not be understood as limiting the present application. Those skilled in the art can understand that various changes, modifications, replacements and variations can be made to these examples without departing from the principles and purposes of the present application, and the scope of the present application is defined by the claims and their equivalents. The experimental consumables, reagents and raw materials used in the present application are easily obtained by those skilled in the art, and can be obtained from commercial channels if not otherwise specified, and the experimental methods not specified in the present application are usually carried out according to conventional conditions or according to the conditions recommended by the manufacturer. In particular, the following examples are only used to illustrate the present application and should not limit the scope of the present application in any way. It should be noted that the experimental conditions and results described in the following examples are only used to illustrate the present application and should not and will not limit the present application as described in detail in the claims.

[0077] Example 1: Isolation, screening and identification of Ruminococcus torques

[0078] 1. Sample source

[0079] Tumor tissues and paired normal intestinal epithelial tissues from colorectal cancer patients were collected, and a total of 582 patient samples were collected. After sample collection, the samples were quickly placed in sterile, low-temperature storage conditions to ensure the activity and integrity of the microorganisms.

[0080] 2. Screening method

[0081] The difference in flora in the tissues was analyzed by 16S rRNA sequencing, and strains with significantly higher abundance in normal tissues were screened. First, the microbial DNA in the samples was extracted and amplified to construct a 16S rRNA gene library, and high-throughput sequencing technology was used for sequencing. Then, bioinformatics analysis methods were used to process the sequencing data and compare the differences in microbial species and abundance in different tissue samples.

[0082] 3. Bacterial identification

[0083] The 16S rRNA gene sequencing of the screened strains was performed, and after comparison, it was identified as Ruminococcus torques. The sequence obtained by sequencing was compared with the sequence in the authoritative database such as NCBI, and the similarity was as high as 99%, confirming that the strain was Ruminococcus torques.

[0084] The 16S rRNA sequence of Ruminococcus torques according to the present application is shown below (SEQ ID NO. 1):

[0085]

[0086] The distribution of the strain in normal intestinal epithelium and tumor tissues is shown in FIG. 1, and the results show that the strain has a high abundance in healthy intestinal epithelial tissue and a significantly reduced abundance in colorectal cancer tissue. Through detection and analysis of a large number of colorectal cancer patient surgical resection samples using 16S rRNA sequencing technology, the distribution difference of the strain in different tissues is clearly presented. In normal intestinal epithelial tissue samples, the relative abundance of the strain can reach 0.6726, while in tumor tissue samples, the relative abundance is only 0.2036, and the difference is statistically significant (P < 0.01). Figure 1

[0087] Example 2 Inhibitory effect of the strain on the growth of subcutaneous tumors in mice

[0088] 1. Model construction

[0089] The colorectal cancer cell line MC38 of mice was inoculated into the axillary of C57BL / 6 mice to construct a subcutaneous tumor model. Healthy C57BL / 6 mice of 4 weeks old and similar weight were selected, and a certain concentration of MC38 cell suspension was injected into the subcutaneous tissue of the axillary of the mice under sterile conditions. When the tumor was formed and the volume reached a certain size, the subsequent experiment was carried out.

[0090] 2. Intervention treatment

[0091] After the tumor was formed, the strain was injected into the tumor of the model mice. The mice were randomly divided into two groups, and 1 x 10 8 CFU (colony forming unit) of the strain was injected into the tumor of each mouse in the experimental group, and an equal amount of sterile normal saline was injected into the tumor of each mouse in the control group.

[0092] 3. Experimental results

[0093] The results are shown in FIG. 2. Compared with the control group, the volume and weight of the subcutaneous tumor of the mice injected with the strain were significantly reduced, indicating that the strain can inhibit tumor growth. The long diameter and short diameter of the tumor were measured regularly with a vernier caliper, the tumor volume was calculated, and the growth curve was drawn. After the experiment was completed, the mice were sacrificed, the tumor was peeled off, and the weight was measured. The results show that the average volume of the tumor of the mice in the control group was 654.4 ± 68.84 mm 3 , and the average weight was 0.883 ± 0.099 g, while the average volume of the tumor of the mice in the experimental group was 474 ± 122.77 mm 3 , and the average weight was 0.708 ± 0.128 g, and the difference was statistically significant (P < 0.05). Figure 2 Example 3 Inhibitory effect of the strain on inflammation-induced colorectal cancer

[0094]

[0095] ​​1. Model construction

[0096] AOM / DSS was used to induce C57BL / 6 mice to establish an inflammation-related colorectal cancer model. The mice were first injected intraperitoneally with AOM (azoxymethane), and then given DSS (dextran sulfate sodium)-containing drinking water one week later, and the cycle was repeated to induce inflammation-related colorectal cancer in the mice.

[0097] 2. Intervention treatment

[0098] The model mice were given gavage treatment with S. ruminantium. The model mice were randomly divided into two groups, and the experimental group was given gavage with 1x10 8 CFU of S. ruminantium per day, and the control group was given gavage with an equal amount of sterile normal saline.

[0099] 3. Experimental results

[0100] The results are shown in Figure 3 Compared with the control group, the number and volume of colon tumors in the intervention group were significantly reduced, indicating that it can inhibit the occurrence of inflammation-induced colorectal cancer. After the experiment, the mice were dissected, the colon tumor was observed, the number of tumors was counted and the volume was measured. The results showed that the average number of colon tumors in the control group was 6.875±1.166, and the average tumor volume was 49.375±11.68 mm 3 , while the average number of tumors in the experimental group was 4.75±1.199, and the average tumor volume was 31.5±6.85 mm 3 , and the difference was statistically significant (P<0.05).

[0101] In addition, the freeze-etching electron microscopy pictures of MC38 cells co-cultured with S. ruminantium are shown in Figure 4 The results show that S. ruminantium can enter the colorectal tumor cells to play a role, directly affecting the tumor microenvironment, and specifically inhibiting the progression of colorectal cancer. After the strain enters the tumor cells, it can regulate the related signaling pathways in the tumor cells, such as affecting the expression of cell proliferation and apoptosis-related genes, thereby inhibiting the growth of tumor cells, and has strong targeting.

[0102] The experimental results of the above examples show that the S. ruminantium provided by the present application has a higher abundance in normal intestinal tissue and has a significant inhibitory effect on colorectal cancer. It can enter tumor cells to function, providing a new strain resource and application direction for the prevention and treatment of colorectal cancer.

Claims

1. The use of *Ruminococcus tortifolius* in the preparation of medicaments for the treatment and / or prevention of colorectal cancer, characterized in that, The sequence of the *Ruminococcus twitchis* is shown in SEQ ID NO.

1.

2. The application according to claim 1, characterized in that, The aforementioned Ruminococcus twistans can inhibit the proliferation of colorectal cancer cells and / or promote apoptosis of colorectal cancer cells.

3. The application according to claim 1, characterized in that, The drug also contains a pharmaceutically acceptable carrier; Optionally, the pharmaceutically acceptable carrier may be a filler, enteric coating material, adhesive, stabilizer, diluent, suspending agent, antibacterial agent, rectal preparation carrier, intratumoral implantation preparation carrier, prebiotic carrier, or inert carrier.

4. The application according to claim 1, characterized in that, The dosage form of the drug is an oral preparation, an injectable preparation, an enteric-coated preparation, a colon-targeted preparation, a rectal preparation, an intratumoral implantation preparation, a lyophilized preparation, a live bacteria capsule preparation, or a microcapsule preparation.

5. A type of Ruminococcus tortifolius with inhibitory activity against colorectal cancer, characterized in that, The sequence of the *Ruminococcus twitchis* is shown in SEQ ID NO.

1.

6. A pharmaceutical composition for treating and / or preventing colorectal cancer, characterized in that, The pharmaceutical composition comprises the Ruminococcus twitche as described in claim 5.

7. A pharmaceutical preparation for treating and / or preventing colorectal cancer, characterized in that, The pharmaceutical preparation comprises the pharmaceutical composition of claim 6; Optionally, the dosage form of the pharmaceutical preparation is an oral preparation, an injectable preparation, an enteric-coated preparation, a colon-targeted preparation, a rectal preparation, an intratumoral implantation preparation, a lyophilized preparation, a live bacteria capsule preparation, or a microcapsule preparation.

8. Any of the following products: (1) A bacterial agent for the treatment and / or prevention of colorectal cancer, characterized in that, The bacterial agent contains the *Ruminococcus tortifolia* as described in claim 5; (2) A food for treating and / or preventing colorectal cancer, characterized in that the food contains the Ruminococcus tortifolius as described in claim 5; (3) A health product for treating and / or preventing colorectal cancer, characterized in that the health product contains the Ruminococcus tortifolia as described in claim 5; (4) A dietary supplement for the treatment and / or prevention of colorectal cancer, characterized in that the dietary supplement contains the Ruminococcus twitche as described in claim 5; (5) A diagnostic product for diagnosing colorectal cancer, characterized in that the diagnostic product comprises a reagent for detecting the abundance of Ruminococcus twitchae as described in claim 5.

9. A method for inhibiting the proliferation of colorectal cancer cells and / or promoting apoptosis of colorectal cancer cells in vitro without therapeutic intent, characterized in that, The method includes contacting the desired cell system with the Ruminococcus twitchae as described in claim 5.

10. Applied to any of the following aspects: (1) The use of the Ruminococcus tortifolia according to claim 5 in the preparation of bacterial agents, foods, health products or dietary supplements for the treatment and / or prevention of colorectal cancer; (2) The use of the reagent for detecting the abundance of Ruminococcus twitchae as described in claim 5 in the preparation of diagnostic products for the diagnosis of colorectal cancer.

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