CN-121518393-B - Method for preparing cell-derived exosomes, product and application thereof

Abstract

The invention discloses a method for preparing cell-derived exosomes, a product and application thereof, wherein the method comprises the following steps: the immune activated programmable cells are co-cultured with a stimulator with immune activation function, and cell-derived exosomes with immune activity are prepared by a natural secretion collection method or an artificial extrusion collection method. The invention has safe and controllable local immune activation, and the immune activation strategy based on exosomes fully overcomes the safety bottleneck of the traditional live bacteria preparation in tumor treatment. By replacing living bacteria with engineering exosomes, not only is the potential risks of systemic inflammatory reaction, infection and the like effectively avoided, but also the local controllability of immune stimulation signals is ensured, the accurate activation and directional regulation and control of immune cells in a tumor area are realized, the tumor microenvironment is remodeled, the anti-tumor effect is enhanced, and the broad-spectrum cross-tumor potential of cross-model verification is enhanced.

Inventors

• Qiao Haishi
• Cong Chunyu
• XIAO ZIYUAN
• Zhang Tiyue
• CAI YUANYUAN

Assignees

• 中国药科大学

Dates

Publication Date

20260508

Application Date

20260113

Claims (7)

1. 1. A method for preparing cell-derived exosomes is characterized by comprising the following steps of co-culturing immune activated programmable cells with an immune activating stimulator, wherein the immune activated programmable cells are macrophages, and the immune activating stimulator is escherichia coli ESCHERICHIA COLI O111:111.
2. 2. A cell-derived exosome prepared by the method of claim 1.
3. 3. Use of the cell-derived exosome of claim 2 in the manufacture of a medicament or pharmaceutical composition for the treatment and/or prevention of breast cancer.
4. 4. Use of the cell-derived exosome and α -PD1 according to claim 2 for the preparation of a pharmaceutical composition for the treatment and/or prevention of breast cancer.
5. 5. A medicament or pharmaceutical composition for the treatment and/or prophylaxis of breast cancer, characterized in that it contains an exosome of cellular origin according to claim 2.
6. 6. The medicament or pharmaceutical composition according to claim 5, wherein the dosage form of the medicament or pharmaceutical composition comprises an injection, a patch, an oral preparation.
7. 7. The medicament or pharmaceutical composition according to claim 5, wherein the pharmaceutical composition further comprises a-PD 1.

Description

Method for preparing cell-derived exosomes, product and application thereof Technical Field The invention relates to a method for preparing an exosome from a cell source, a product and application thereof, and belongs to the field of exosome application. Background The tumor is a serious disease seriously threatening human health, the treatment of the tumor still faces a plurality of challenges, and the malignant tumor has the characteristics of high heterogeneity, easy recurrence and metastasis, tolerance to conventional treatment and the like, so that even if the existing treatment means are continuously advanced, the curative effect is still not ideal for a plurality of advanced patients, and the overall survival rate needs to be improved. With the rapid development of immunology, immunotherapy provides a new direction for tumor treatment, and this therapeutic approach by activating or enhancing the recognition of human autoimmune system and attacking tumor cells has shown remarkable therapeutic effects in various solid tumors. Specifically, it can enhance anti-tumor immune response by targeting immune checkpoint molecules such as PD-1, PD-L1, CTLA-4 to relieve tumor immunosuppression, or directly expanding tumor-specific killer T cells by means of adoptive immune strategies such as CAR-T cell therapy. The immunotherapy shows remarkable curative effects in various solid tumors such as melanoma, lung cancer, lymphoma, glioma and the like and blood system tumors, but the clinical application still faces key bottlenecks that on one hand, physical barriers of tumor tissues (such as compact matrixes of solid tumors, blood brain barriers related to brain tumors and the like) severely limit therapeutic drugs and immune effector cells to enter tumor parts, so that local immune activation is insufficient, and on the other hand, immunosuppressive microenvironments which almost exist in all tumors are characterized in that the immunosuppressive microenvironments show depletion of immune cell functions (such as high expression of PD-1, TIM-3 and other depletion markers of CD8 + T cells), enrichment of suppressive immune cells (such as regulatory T cells and M2 type macrophages), infiltration of effector lymphocytes and the like, and the two together lead to poor immune therapeutic effects of partial tumors. Under the background, the new concept of immunity training provides a new idea for solving the problems, and the core of the new concept is that the innate immune cells can form a long-term immune memory-like state through epigenetic regulation modes such as histone modification, DNA methylation and the like after being stimulated by specific stimulation, and the new concept is accompanied by metabolic recombination characteristics such as glycolysis enhancement and the like, so that the immune response can be started faster and stronger pro-inflammatory factors (such as IL-6 and TNF-alpha) secretion and phagocytic killing activity can be generated when the new immune cell is stimulated by tumor antigens and the like later. BCG vaccine (BCG) is used as classical training immunity inducer, can activate innate immune cells such as macrophages and dendritic cells by combining cell wall component lipoarabinomannan and the like with pattern recognition receptors such as TLR2, TLR4 and the like on the surface of immune cells, can directly enhance the phagocytic clearance capacity of the innate immune cells to tumor cells, can promote the maturation of the dendritic cells and present tumor antigens, and further activates CD4 + auxiliary T cells and CD8 + cytotoxic T cells to start adaptive immune response, and has potential value in tumor immunotherapy. However, BCG has obvious limitations in clinical application, such as possible systemic inflammatory reaction and other systemic side effects caused by the BCG as a live bacteria preparation, difficult accurate control of in vivo immune activation intensity and time, easy generation of excessive or insufficient immune response, difficult crossing of blood brain barrier to act on intracranial tumor sites, incapacity of effectively activating intracranial local immunity, possibility of weakening treatment effect due to immune tolerance caused by long-term use, and serious limitation of application prospects of training immune inducers such as BCG in tumor immunotherapy. And a novel exosome-based therapeutic strategy provides a breakthrough solution to overcome the limitations of traditional training immunotherapy. Compared with the therapy based on living bacteria, the exosome has multiple remarkable advantages that nanoscale dimensions and engineering surface characteristics (such as modified tumor targeting ligands, blood brain barrier targeting peptides and the like) endow the exosome with good tissue penetrating capacity, the exosome is beneficial to crossing over various tumor physical barriers such as solid tumor matrixes, blood brain barriers and the like, the accur