106 results about "Gamma delta T cell" patented technology

The invention provides a gamma delta T cell preparation as well as a preparation method and application thereof, and belongs to the technical field of T cells. The liposome is prepared from the following raw materials in parts by weight: 15-20 parts of modified gamma delta T cell liposome, 1-3 parts of cell stimulating factors and 4-7 parts of culture medium freeze-dried powder, the modified gamma delta T cell lipidosome is prepared by embedding gamma delta T cells through lipidosome, coupling with acetylenic bond modified chitosan, and further mixing with MFAP4 protein and b4GALT1 protein. The culture medium freeze-dried powder is prepared by freeze-drying the culture medium in the engineering culture process of gamma delta T cells, and the cell stimulating factors are resveratrol and quercitrin. According to the gamma delta T cell preparation prepared by the invention, the survival ability and resistance of gamma delta T cells are improved, the anti-tumor activity of the gamma delta T cells is improved, and the gamma delta T cell preparation has relatively good antioxidant and anti-inflammatory effects, reduces the inhibition of inflammation on an immune system and has a very good treatment effect on pancreatic ductal adenocarcinoma.

The invention relates to the technical field of biological medicine, in particular to a cell culture medium and a preparation method of gamma delta T cells. The cell culture medium is a culture medium A or a culture medium B. The culture medium A comprises a serum-free culture medium, 1-100 [mu] M of zoledronic acid, 50-500 U / mL of IL-2, 1-5 [mu] M of imiquimod, 1-20 ng / mL of IL-15, 1-50 ng / mL of IL-1b, 10-100 ng / mL of IL-7 and 2-10% (v / v) of inactivated plasma or a serum substitute. The culture medium B comprises a serum-free culture medium, 100 to 2000 U / mL of IL-2, 1 to 50 ng / mL of IL-15, 1 to 10 [mu] M of zoledronic acid and 2 to 10% (v / v) of inactivated plasma or a serum substitute. By adopting the cell culture medium, a large number of gamma delta T cells can be rapidly obtained.

The present disclosure provides genetically modified iPSC-derived γδT cells and their precursors. A double genomic disruption in the suppressor of cytokine signaling 1 (SOCS1) gene and the cytokine-inducible sh2-containing protein (CISH) gene are provided, as is a triple genomic disruption in genes for SOCS1, CISH, and Bcl-2 interacting mediator of cell death (BIM), as is a quadruple genomic disruption in genes for SOCS1, CISH, BIM, and cell surface death receptor (FAS), as is a quintuple genomic disruption in genes for SOCS1, CISH, BIM, β-2-Microglobulin (B2M), and class II transactivator (CITTA), as is a sextuple genomic disruption in genes for SOCS1, CISH, BIM, B2M, CITTA, and FAS. Also provided is genetically modified iPSC-derived γδT cells and their precursors with improved proliferation and tumor killing activity. Also provided are genetically modified iPSC-derived γδT cells and their precursors further comprising CD19 CAR. The present disclosure further provides methods making and using such cells, as well as gene editing systems.

The invention discloses a gamma delta T cell efficient amplification method and application, and belongs to the technical field of cell biology. The amplification method comprises the following steps: separating PBMC (peripheral blood mononuclear cells) by adopting a density gradient centrifugation method, carrying out initial culture through a polylysine coated container, carrying out three-stage dynamic stimulation, combining with an X-VIVO 15 culture medium of 5-8% autoserum, and finally carrying out anti-gamma delta TCR magnetic bead separation to obtain high-purity cells. Through collaborative optimization of stepped factor combination, staged container coating and a low-serum system, the amplification multiple of the gamma delta T cells reaches 150-180 times, the purity is larger than or equal to 90% after purification, the cytotoxicity and the survival ability are remarkably improved, the gamma delta T cells can be efficiently used for immunotherapy of tumors and infectious diseases, and stable technical support is provided for clinical transformation of the gamma delta T cells.

The present disclosure provides genetically modified iPSC-derived γδT cells and their precursors. A double genomic disruption in the suppressor of cytokine signaling 1 (SOCS1) gene and the cytokine-inducible sh2-containing protein (CISH) gene are provided, as is a triple genomic disruption in genes for SOCS1, CISH, and Bcl-2 interacting mediator of cell death (BIM), as is a quadruple genomic disruption in genes for SOCS1, CISH, BIM, and cell surface death receptor (FAS), as is a quintuple genomic disruption in genes for SOCS1, CISH, BIM, β-2-Microglobulin (B2M), and class II transactivator (CIITA), as is a sextuple genomic disruption in genes for SOCS1, CISH, BIM, B2M, CIITA, and FAS. Also provided is genetically modified iPSC-derived γδT cells and their precursors with improved proliferation and tumor killing activity. Also provided are genetically modified iPSC-derived γδT cells and their precursors further comprising CD19 CAR. The present disclosure further provides methods making and using such cells, as well as gene editing systems.

The invention provides a culture medium for gamma delta T cells and a culture method of the gamma delta T cells, and belongs to the technical field of cell culture. According to the culture medium of the gamma delta T cells, OpTmizer serves as a basic culture medium, and the OpTmizer culture medium further contains IL-2 with the concentration being 150-250 IU / mL, IL-6 with the concentration being 40-60 [mu] mol / L, vitamin C with the concentration being 40-60 [mu] mol / L, vitamin B3 with the concentration being 8-12 [mu] mol / L and coenzyme Q10 with the concentration being 3-7 [mu] mol / L. The prepared gamma delta T cell culture medium is used for culturing gamma delta T cells with improved killing activity, and the killing rate of the cultured gamma delta T cells on A549 cells and HepG2 cells can be remarkably improved.

The invention provides a gamma delta T cell culture method and a culture medium, and belongs to the technical field of T cell culture. Modified nanoparticles, zoledronic acid, vitamin C and glutamine are added into a 1640 culture solution, the concentration of the modified nanoparticles ranges from 0.5 g / L to 1 g / L, the concentration of the zoledronic acid ranges from 0.5 mu mol / L to 1.5 mu mol / L, the concentration of the vitamin C ranges from 200 mu mol / L to 500 mu mol / L, and the concentration of the glutamine ranges from 0.1 mmol / L to 0.2 mmol / L. The prepared culture medium can efficiently stimulate amplification of the gamma delta T cells and improve the killing activity of the gamma delta T cells, meanwhile, the survival rate of the gamma delta T cells is also improved, and meanwhile, the culture method of the gamma delta T cells is simple, low in cost and wide in application prospect.

Disclosed herein are methods of stimulating and expanding polyclonal gamma delta T cells (GDTCs) in vitro. More specifically, anti-gamma delta T cell receptor (GDTCR) antibody is used in combination with anti-CD28 antibody and one or more of IL-2, IL-7, and IL-15 to efficiently expand GDTCs in vitro.

The application discloses a preparation method of CCR2+ gamma delta T cells with anti-tumor activity and migration ability, and belongs to the technical field of cells; the method comprises the following steps: step (1), PBMC of peripheral blood of a healthy person is extracted, inoculated into complete culture medium, and added with zoledronic acid, IL-15 and metformin, and then is cultured in a T25 culture bottle; step (2), complete culture medium is supplemented, the cell concentration is adjusted, and then the cells are transferred into a T75 culture bottle for culture; step (3), complete culture medium is supplemented, the cell concentration is adjusted, glucocorticoid and nicotinamide are added, and then the cells are transferred into a T150 culture bottle for culture; step (4), cells are collected at D6, rinsed with normal saline, resuspended with complete culture medium, the cell concentration is adjusted, and then the cells are inoculated into a T300 culture bottle for culture; and step (5), complete culture medium is supplemented every two days from D8 to D16, the cell concentration is maintained, and then the cells are harvested at D16 to obtain CCR2+ gamma delta T cells.

The present disclosure provides, among other things, a method of isolating and expanding gamma delta (γδ) T cells, wherein the method comprises: (a) isolating non-hematopoeitic tissue by biopsy or explant, and (b) culturing the isolated non-hematopoeitic tissue in the absence of a three-dimensional scaffold or grid, thereby expanding and isolating gamma delta (γδ) T cells.

Disclosed herein are compositions and methods of treating a subject with cancer. The compositions and methods utilize immunoresponsive cells to effect killing of tumor cells.

Aspects of the disclosure include methods of treating an autoimmune disease, including reducing the need for chronic immunosuppression, and for effectuating immune reset and / or producing a naive B cell repertoire in a subject in need thereof, the methods comprising administering a therapeutically effective amount of anti-CD20 yδ T cells to the subject, wherein the anti-CD20 yδ T cells express a chimeric antigen receptor (CAR) comprising a binding domain that specifically binds to CD20.

Compositions and methods for stimulating the proliferation and cytotoxicity of γδ T cells are described. Therapeutic compositions and methods using the expanded and stimulated γδ T cells are described.

The invention belongs to the field of medicines, and relates to a gamma delta T cell containing a chimeric antigen receptor EGFR-CCR6-E27CAR. The chimeric antigen receptor comprises a single-chain antibody ScFv of a targeted EGFR (epidermal growth factor receptor), a CD8 hinge region, a CD8 transmembrane region, 4-1BB, a CD3 zeta chain, CCR6 and a secreting type ScFv of a targeted PD-1. The invention also comprises a protein and a nucleic acid sequence of the chimeric antigen receptor, and a preparation method and application of the vector, the protein, the nucleic acid and the cell. In an in-vitro cell model and an in-vivo mouse, the CAR-gamma delta T cell disclosed by the invention shows relatively strong tumor cell killing ability, can remarkably control in-vivo growth of tumors and remarkably prolong the lifetime of the mouse, and lays a foundation for the development of drugs for long-term control of in-vivo tumors of patients.

The application provides an immune cell culture kit, a culture method and application of gamma delta T cells, and the kit comprises a basic culture medium, a recombinant antibody, a cytokine and a cell proliferation promoting additive, the basic culture medium comprises an RPMI-1640 culture medium, and the cell proliferation promoting additive comprises 1-thioglycerol, taurine, beta-cyclodextrin, thymidine, alpha-lipoic acid, beta-mercaptoethanol and bisphosphonate. The gamma delta T cells obtained by the culture kit have high purity and activity, and have good tumor killing activity. The gamma delta T cells obtained by the method have a high positive rate of 100% of perforin expression.

Disclosed herein is a method for enhancing adoptively transferred autologous or allogeneic immune effector T-cells (including gamma delta T cells (γδ-T cells)) by targeting the thioredoxin (TRX)-interacting protein (TXNIP). In addition, disclosed herein are cDNA sequences for the co-expression of immune receptors (CARs, TCRs, etc) in combination with guide RNAs, and / or artificial or natural microRNAs, and / or shRNAs targeting TXNIP.

The present invention relates to a method for treating cancer, comprising administering a composition comprising γδ T cells treated with an inhibitor of XBP1 gene expression or an inhibitor of XBP1 protein expression or activity. The γδ T cells, in which the XBP1 gene is deficient or the XBP1 protein activity is inhibited, exhibit enhanced antitumor activity in a tumor microenvironment characterized by endoplasmic reticulum (ER) stress, as compared to unmodified γδ T cells. Accordingly, the modified γδ T cells are effective for use in cancer treatment.

The invention includes compositions comprising a chimeric antigen receptor (CAR) specific for an GD T Cell receptor (anti-GD TCR CAR), vectors comprising the same, compositions comprising anti-GD TCR CAR vectors packaged in viral particles, and recombinant T cells or other effector cells comprising the anti-GD TCR CAR of the invention. The invention also includes methods of making a genetically modified T cell expressing an anti-GD TCR CAR wherein the expressed CAR comprises an extracellular domain that binds to GD T cells or to cells expressing a GD TCR.

The application provides a kind of gamma delta T cell culture method and culture medium, belong to T cell culture technical field.In 1640 culture solution is added modified nanoparticle, zoledronic acid, vitamin C, glutamine, wherein, the concentration of the modified nanoparticle is 0.5-1g / L, the concentration of the zoledronic acid is 0.5-1.5 μmol / L, the concentration of the vitamin C is 200-500 μmol / L, the concentration of the glutamine is 0.1-0.2 mmol / L.The culture medium prepared by the application can efficiently stimulate gamma delta T cell expansion and improve its killing activity, and the survival rate of gamma delta T cells is also improved, and the culture method of gamma delta T cells is simple, the cost is low, and has broad application prospect.

The application provides a kind of gamma delta T cell preparation and its preparation method and application, it is related to the field of biotechnology.The composition provided by the application includes: coenzyme Q10, vitamin C, human blood albumin, trehalose, sodium pyruvate and amifostine.Using the composition and related preparation provided by the application can effectively improve the killing ability of gamma delta T cells and maintain the function of gamma delta T cells, and can effectively inhibit tumor growth.

The invention provides a gamma delta T cell, a proliferation culture method thereof and application of the gamma delta T cell as a medical preparation, and belongs to the technical field of medical preparations. The proliferation culture method for gamma delta T cells provided by the invention comprises the following steps: (1) separating mononuclear cells in peripheral blood, inoculating the mononuclear cells into a culture medium containing a chemical stimulant and cytokines, and culturing the mononuclear cells; (2) supplementing cell factors on the 3th day of culture, and continuing to culture until the multiplication culture is finished; the chemical stimulant is isopentenyl pyrophosphate, and the cell factor is IL-2 (Interleukin-2). According to the proliferation culture method of the gamma delta T cells, provided by the invention, the proliferation efficiency of the gamma delta T cells and the killing activity of the gamma delta T cells on lung cancer cells are remarkably improved, and when culture is finished, the proliferation multiple of the gamma delta T cells is 10000 times, and the killing efficiency of the gamma delta T cells on the lung cancer cells is 70%, which are remarkably higher than those in the prior art.

To provide a method for producing a γδT cell population which expresses functional TCR or CAR by preparing large quantities of γδT cells with high purity and efficiently introducing a foreign gene encoding TCR or CAR into the obtained γδT cells.SOLUTION: The present invention provides a method for efficiently producing highly pure γδT cells introduced with foreign genes by culturing in the presence of IL-7 and IL-15 after stimulating the γδT cells with a bisphosphonic acid ester derivative, followed by gene introduction. When a T cell receptor (TCR) or a chimeric antigen receptor (CAR) is introduced as the foreign gene, the γδT cells in which TCR and CAR are functionally expressed can be obtained.SELECTED DRAWING: None

The present invention relates to a method for producing gamma delta (γδ) T cells. The present invention also relates to the cells obtained by said method and said cells for their use as a medicament and in an immunotherapy, preferably in the treatment or prevention of a cancer.

The present invention relates to bispecific antibodies that target the BTN3A receptor and block the PD-1 / PD-L1 inhibition axis. The invention also relates to bispecific antibodies capable of targeting immune cells, in particular gamma delta T cells, to tumor cells expressing PD-L1. The invention also relates to methods of treating cancer and pharmaceutical compositions comprising said bispecific antibodies.

The invention discloses a culture medium for in-vitro amplification and activation of gamma delta T cells, a culture method and application of the culture medium, and belongs to the technical field of in-vitro culture of immune cells. The culture medium comprises zoledronic acid, interleukin-2 (IL-2), wnt / beta-catenin signaling pathway activators (GSK-3alpha and GSK-3beta specific inhibitors), particularly 6-bromo indirubin-3 '-acetoxime (Bio-acetoxime), peripheral blood mononuclear cells (PBMC) can be used as a starting material, gamma delta T cells can be separated from the PBMC through amplification for 14 days, high-purity gamma delta T cells can be efficiently and repeatedly generated, and high-purity gamma delta T cells can be obtained. The yield of the gamma delta T cell can be obviously improved, the obtained gamma delta T cell has the functions of a T cell, an NK cell and an antigen presenting cell, and the anti-tumor efficiency is also obviously improved.

To provide compositions and methods for treatment of hematological tumors with engineered or non-engineered γδ-T cells.SOLUTION: In some embodiments, γδ-T cells comprise a chimeric antigen receptor (CAR) construct. The CAR construct can contain an anti-CD20 binding domain or anti-B cell maturation antigen (BCMA) binding domain, a CD8 hinge and transmembrane domain, a costimulatory domain, a CD3ζ signalling domain, a combination thereof, or all thereof. The CAR construct can contain a domain encoding for a secreted common gamma chain cytokine such as an sIL15 domain.SELECTED DRAWING: None

The invention belongs to the technical field of cytobiology and immune cell therapy, and particularly relates to a gamma delta T cell serum-free additive culture medium composition with definite components and application and a product thereof. The serum-additive-free culture medium composition comprises a serum-additive-free culture medium A, a serum-free culture addition reagent B, a serum-additive-free culture medium C, a serum-additive-free culture medium D and a serum-additive-free culture medium E, the main components comprise ethanolamine, sodium selenite, recombinant human albumin, IL-15 and IL-2. According to the gamma delta T cell serum-free additive culture medium composition provided by the invention, the components are clear, the serum-free culture additives without animal-derived components can avoid potential safety hazards caused by the animal-derived components, the amplification efficiency is improved, the cell high activity is maintained, the GMP standard is met, and the gamma delta T cell serum-free additive culture medium composition is suitable for large-scale production and clinical transformation.

The invention discloses a composite immune cell co-culture system, which comprises an immune cell system and a cell factor system, the immune cell system comprises the following components in percentage by weight: 10%-60% of CD8 + T cells, 5%-35% of NK cells, 5%-20% of gamma delta T cells, 10%-30% of CD4 + Th1 cells, 3%-20% of DC cells, and 2%-20% of B cells; the cell factor system comprises IL-2, IL-7, IL-12, IL-15, IL-18, TNF (Tumor Necrosis Factor)-alpha and IL-1beta. According to the invention, through differentiated ratio design of a starting period, an amplification period and a memory induction period, activation, proliferation and differentiation requirements of immune cells in each stage are accurately matched, the problem that static ratio cannot give consideration to multi-stage functions is solved, and the overall activity of a system is remarkably improved.

The application belongs to the technical field of biotechnology, and particularly relates to a staged precise regulation in vitro expansion culture method of high-activity gamma delta T cells. In the synergistic activation stage, the complex activator containing zoledronic acid, IL-2, IL-15, IL-12 and IL-21 is used to significantly improve the early activation level of the gamma delta T cells; in the dynamic expansion stage, the concentration of metformin is adjusted step by step to support the continuous and efficient proliferation of the cells; in the gradient maturation stage, the combination system of N-acetyl cysteine, TWS119 and DL-alpha tocopheryl acetate is introduced to help maintain the functional activity and antioxidant state of the cells in the later culture stage. The gamma delta T cells with a purity higher than 90% and an expansion multiple more than 100 times can be obtained by using the method to culture for 10 days, and the cells exhibit significant killing activity to tumor cells, are suitable for the adoptive immunotherapy of tumor and infectious diseases, and have good clinical application prospects.

The present invention relates to methods for introducing nucleic acid into innate-like lymphocytes, including gamma delta T cells, with a lentiviral vector; populations or compositions of genetically engineered innate-like lymphocytes; and methods for treating or preventing disease.