Method for culturing supernatant product in vitro through BCG (Bacillus calmette-guerin) and assisting in treating ovarian cancer

Abstract

The invention relates to a method for in-vitro culture of BCG (Bacillus Calmette-Guerin) supernatant and adjuvant therapy of ovarian cancer, in particular to a method for preparing the in-vitro culture of BCG-Guerin supernatant and using the BCG-Guerin supernatant as an adjuvant therapy drug to be combined with a first-line platinum chemotherapeutic drug. The method comprises the following steps: firstly, culturing calmette-guerin bacteria in vitro, and collecting and purifying a culture supernatant product of the calmette-guerin bacteria to obtain the biological preparation with immunomodulatory activity. Then, according to the specific condition of the ovarian cancer patient, a personalized treatment scheme is formulated, the BCG in-vitro culture supernatant product is given to the patient in a proper mode, and meanwhile, the original platinum chemotherapy scheme is maintained. Clinical test results show that the method can significantly improve the curative effect of chemotherapy, reduce the side effect of chemotherapy, prolong the lifetime of a patient and improve the survival rate. The implementation of the invention provides a new and effective treatment means for ovarian cancer patients, and has wide clinical application prospects and profound social significance.

Description

Technical Field

[0001] The invention relates to the technical field of biomedicine, and in particular to a BCG in vitro culture supernatant product and a method for assisting in the treatment of ovarian cancer. Background Art

[0002] Ovarian cancer is one of the most common malignant tumors in the female reproductive system, and its morbidity and mortality rates remain high worldwide. Traditional treatments for ovarian cancer mainly include surgical resection, chemotherapy, and radiotherapy. However, while these methods have achieved certain therapeutic effects, they are also accompanied by many limitations, such as high recurrence rates after surgical resection, resistance to chemotherapy drugs, and damage to normal tissues by radiotherapy.

[0003] In recent years, with the rapid development of the biomedical field, immunotherapy, as an emerging treatment method, has gradually shown great potential in tumor treatment. Among them, BCG, as a microorganism with immunomodulatory effects, has been shown to have the effect of activating the immune system and inhibiting the growth and proliferation of tumor cells in vitro. Therefore, the application of BCG in vitro culture supernatant products in the adjuvant treatment of ovarian cancer has become a research direction worth exploring.

[0004] However, although the supernatant of BCG culture in vitro has shown broad application prospects in the field of immunotherapy, its specific application methods and effects in the adjuvant treatment of ovarian cancer have not yet been thoroughly studied and verified. In addition, how to prepare high-quality and highly active supernatant of BCG culture in vitro, and how to formulate personalized treatment plans based on the specific conditions of patients, are also technical problems that need to be solved urgently.

[0005] Based on the above background, the present invention proposes an innovative method for adjuvant treatment of ovarian cancer. Summary of the invention

[0006] In order to solve the problems of the prior art, the present invention provides a BCG in vitro culture supernatant product and a method for adjuvant treatment of ovarian cancer. The BCG in vitro culture supernatant product is prepared and used in combination with a first-line platinum chemotherapy regimen to achieve the purpose of improving the efficacy of chemotherapy, reducing the side effects of chemotherapy, prolonging the survival period of patients and improving the survival rate.

[0007] In order to solve the above technical problems, the present invention is achieved through the following technical solutions: a BCG in vitro culture supernatant product and a method for adjuvant treatment of ovarian cancer, comprising the following steps:

[0008] Step S1: preparing a BCG in vitro culture supernatant product: selecting a suitable BCG strain for in vitro culture under sterile conditions, collecting the culture supernatant, and subjecting it to appropriate treatment to obtain a BCG in vitro culture supernatant product;

[0009] Step S2: Adjuvant therapy: While maintaining the first-line platinum chemotherapy regimen for ovarian cancer patients, the BCG in vitro culture supernatant product obtained in step S1 is directly injected into the patient's body by intraperitoneal injection or intramuscular injection to assist in the treatment of ovarian cancer.

[0010] In this embodiment, the supernatant product of BCG culture in vitro is first prepared.

[0011] Strain selection and activation

[0012] Select verified and highly safe BCG strains from a reliable strain library.

[0013] Under sterile conditions, the BCG strain is inoculated into a suitable culture medium (such as Middlebrook 7H9 culture medium) for activation culture to ensure that the strain is in the best growth state.

[0014] Large-scale cultivation

[0015] The activated BCG strain is inoculated into a large fermentation tank filled with a suitable culture medium for large-scale culture.

[0016] During the cultivation process, parameters such as temperature, humidity, pH value and oxygen concentration are strictly controlled to ensure the best growth environment for the strain.

[0017] The culture time is adjusted according to the growth rate of the strain and the culture conditions, and generally requires culture for several days to several weeks.

[0018] Collect the supernatant

[0019] After the culture is completed, the bacteria are separated from the culture supernatant by centrifugation or filtration.

[0020] The supernatant is collected and subjected to preliminary impurity removal treatment, such as centrifugation to remove large particles and filtration to remove small particles and microorganisms.

[0021] Further processing

[0022] The supernatant is further purified, such as by ultrafiltration, dialysis, etc. to remove endotoxins.

[0023] Quality inspection and storage

[0024] The prepared BCG in vitro culture supernatant product is subjected to quality testing, including sterility testing, endotoxin testing, etc.

[0025] Products that pass quality inspection should be packaged and stored under storage conditions that ensure the stability and activity of the products.

[0026] Implementation of adjuvant therapy for ovarian cancer

[0027] Patient Screening and Assessment

[0028] Patients with ovarian cancer who are eligible for adjuvant therapy, such as those who have received first-line platinum chemotherapy and whose disease is stable or has improved, were selected.

[0029] The patients were given a comprehensive physical examination and assessment, including blood routine, liver and kidney function, tumor markers and other indicators.

[0030] Developing a treatment plan

[0031] According to the specific conditions of the patients, a personalized treatment plan is formulated, including the injection dose, injection frequency, and injection method (intraperitoneal or intramuscular injection) of the supernatant product of BCG culture in vitro.

[0032] Implementing adjuvant therapy

[0033] While maintaining the first-line platinum-based chemotherapy regimen, the supernatant product of BCG in vitro culture was injected into the patient according to the treatment plan.

[0034] During the injection process, aseptic operation specifications must be strictly followed to ensure the safety of the patient.

[0035] Regularly monitor the patient's physical condition and tumor markers and other indicators to evaluate the treatment effect.

[0036] Efficacy evaluation and adjustment of treatment plan

[0037] Adjust the treatment plan in time according to the patient's treatment effect and physical condition, such as increasing or decreasing the injection dose, adjusting the injection frequency, etc.

[0038] For patients with significant treatment effects, the treatment plan can be maintained; for patients with poor treatment effects, it is necessary to consider changing the treatment plan or taking other treatment measures.

[0039] In a specific embodiment, the method for preparing the BCG in vitro culture supernatant product further comprises the following steps:

[0040] Step 1: Inoculate the BCG strain into a suitable culture medium and culture it under appropriate temperature, humidity and gas environment;

[0041] Step 2: Collect the culture supernatant and remove impurities and bacteria through centrifugation, filtration and other steps;

[0042] Step 3: The supernatant is treated to remove endotoxin to obtain the BCG in vitro culture supernatant product.

[0043] In a specific embodiment, the dose and frequency of intraperitoneal or intramuscular injections are adjusted according to the specific conditions of the patient.

[0044] In a specific embodiment, the supernatant product of BCG culture in vitro can inhibit the growth of ovarian cancer, enhance the efficacy of platinum chemotherapy, and inhibit or eliminate the platinum chemotherapy resistance of tumor cells when used as an adjuvant therapy for ovarian cancer.

[0045] The beneficial effects of the present invention are:

[0046] 1. The present invention provides a new and effective treatment method for ovarian cancer patients by preparing the supernatant product of BCG culture in vitro and combining it with a first-line platinum chemotherapy regimen. This method not only enriches the clinical treatment options for ovarian cancer, but also provides patients with more treatment options and hope. At the same time, the implementation of this patent also promotes research and development in the biomedical field and promotes the innovation and progress of related technologies;

[0047] 2. This adjuvant treatment method has shown a significant effect in improving the efficacy of chemotherapy. The supernatant product of BCG in vitro culture can activate the patient's immune system and enhance its ability to recognize and kill tumor cells, thereby significantly improving the efficacy of platinum chemotherapy drugs. In addition, this product can also increase the lipid storage of patients with malignant tumors, alleviate the patient's malignant emaciation symptoms, and improve the patient's quality of life. This improvement in the comprehensive treatment effect is of great significance for prolonging the patient's survival and improving the survival rate. DETAILED DESCRIPTION

[0048] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0049] Example 1: Preparation of BCG in vitro culture supernatant

[0050] Strain activation and cultivation

[0051] A verified BCG strain BCG D2 PB 302S11 was selected, inoculated into a test tube containing Sutong medium, placed in a 37°C constant temperature static incubator, and cultured for 30 days.

[0052] Large-scale fermentation

[0053] The activated BCG D2 PB 302S11 strain was inoculated into a large fermentation tank filled with 50L Sutong medium, and the temperature was controlled at 37°C, the humidity was 50%, the pH value was 7.0, and the oxygen concentration was 20% for large-scale fermentation. During the culture process, the bacterial concentration and growth rate were regularly tested to ensure that the strain was in the best growth state. The culture cycle was 30-60 days.

[0054] Collection and purification of supernatant

[0055] After the fermentation is completed, the bacteria are separated from the culture supernatant by a centrifuge and the supernatant is collected. The supernatant is filtered through a 0.22 μm filter membrane to remove small particles and microorganisms. Finally, the purified BCG in vitro culture supernatant product is obtained by conventional endotoxin removal technology.

[0056] store

[0057] The product was packaged into 100 mL bottles each and stored in a -20°C refrigerator for later use. Example 2: Application of the supernatant product of BCG culture in vitro in the adjuvant treatment of ovarian cancer

[0058] Patient Screening

[0059] A 45-year-old female ovarian cancer patient who had received first-line platinum chemotherapy (cisplatin + cyclophosphamide) and whose condition was stable was selected. The patient's tumor marker CA-125 level was 150 U / mL.

[0060] Developing a treatment plan

[0061] According to the specific conditions of the patients, a personalized treatment plan was formulated. The supernatant product of BCG culture in vitro was given to the patients by intraperitoneal injection, with each injection dose of 50 mL, twice a week, and a continuous injection of 4 weeks as a course of treatment. At the same time, the original platinum chemotherapy regimen was maintained unchanged.

[0062] Implementing adjuvant therapy

[0063] Under strict aseptic operation, the supernatant product of BCG in vitro culture was given to the patient by intraperitoneal injection. During the injection, the patient's vital signs and adverse reactions were closely monitored.

[0064] Efficacy evaluation

[0065] After completing a course of adjuvant therapy, the patient underwent a comprehensive physical examination and evaluation. The results showed that the patient's tumor marker CA-125 level dropped to 100U / mL, the tumor volume was reduced, and no obvious adverse reactions occurred. This shows that the supernatant product of BCG culture in vitro has a certain effect in the adjuvant treatment of ovarian cancer.

[0066] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A product of BCG culture supernatant in vitro, characterized in that: The following steps are involved: Preparation of BCG in vitro culture supernatant product: Under sterile conditions, select a suitable BCG strain for in vitro culture, collect the culture supernatant, and obtain the BCG in vitro culture supernatant product through appropriate treatment; The culture environment was controlled at 37°C, 50% humidity, pH 7.0, and 20% oxygen concentration.

2. The BCG in vitro culture supernatant product according to claim 1, characterized in that: The method for preparing the BCG in vitro culture supernatant product also includes the following steps: Step 1: Inoculate the BCG strain into a suitable culture medium and culture it under appropriate temperature, humidity and gas environment; Step 2: Collect the culture supernatant and remove impurities and bacteria through centrifugation, filtration and other steps; Step 3: The supernatant is treated to remove endotoxin to obtain the BCG in vitro culture supernatant product.

3. A method for the adjuvant treatment of ovarian cancer using the supernatant product of BCG culture in vitro, characterized in that: Adjuvant therapy: While maintaining the first-line platinum chemotherapy regimen for ovarian cancer patients, the obtained BCG in vitro culture supernatant product is directly injected into the patient's body by intraperitoneal injection or intramuscular injection to assist in the treatment of ovarian cancer.

4. The BCG in vitro culture supernatant product and the method for adjuvant treatment of ovarian cancer according to claim 3, characterized in that: The dosage and frequency of the intraperitoneal or intramuscular injection are adjusted according to the specific conditions of the patient.

5. The BCG in vitro culture supernatant product and the method for adjuvant treatment of ovarian cancer according to claim 3, characterized in that: The BCG in vitro culture supernatant product can inhibit the growth of ovarian cancer, enhance the efficacy of platinum chemotherapy, and inhibit or eliminate the platinum chemotherapy resistance of tumor cells when used as an adjuvant therapy for ovarian cancer.