42 results about "NKG2D" patented technology

The application discloses a kind of nanobody specifically targeting human and murine NKG2D protein and its preparation method and application.The nanobody or antigen-binding fragment thereof of the application has three complementarity determining regions CDR1, CDR2 and CDR3;Wherein the amino acid sequences of CDR1, CDR2, CDR3 are as shown in SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4 respectively.The nanobody can simultaneously bind hNKG2D and mNKG2D with high affinity, and the bispecific nanobody E5 / 2-B12 based on the nanobody can specifically bind CEACAM5 positive tumor cells and NKG2D overexpression cells, and effectively activate NK cells, which can significantly inhibit tumor growth and prolong survival in various tumor models, and has good application prospect in tumor immunotherapy.

The invention includes a chimeric antigen receptor (CAR) specific for a natural-killer group 2, member D ligand (NKG2DL), compositions comprising the CAR, polynucleotides encoding the CAR, vectors comprising a polynucleotide encoding the CAR, and recombinant T cells comprising the CAR. The invention also includes methods of making a genetically modified T cell expressing a NKG2D-CAR wherein the expressed CAR comprises a NKG2D extracellular domain. The present invention also relates generally to the use of T cells engineered to express a CAR to treat cancer associated with NKG2D.

The invention provides application of CAR-NK (chimeric antigen receptor-natural killer) cells in preparation of drugs for treating autoimmune diseases, and the CAR-NK cells express a chimeric antigen receptor and a fusion protein of a targeted NKG2D ligand; the fusion protein comprises an IL-15R alpha and an IL-15, and the IL-15 and the IL-15R alpha are connected with each other. According to the CAR-NK cell disclosed by the invention, the specific recognition of MICA / B molecules on the surface of an over-activated T / B cell of an SLE patient is realized by highly expressing an NKG2D activated receptor, and the durability of an intracellular signal is enhanced so as to resist cell depletion; meanwhile, the super IL-15 fusion protein is expressed to prolong the in-vivo survival and function duration time, and the effect of treating the SLE is improved through the cooperation of the dual mechanisms.

Provided is the use of NKG2D-CD3 and NKG2D-CD16 cell adapter molecules in the removal of aging cells and the treatment of aging-cell-accumulation-related diseases, wherein the cell adapter molecules comprise an NKG2D extracellular domain, a connection segment, and a CD3 or CD16 binding domain. The death of aging cells having high expression of NKG2D ligand is efficiently induced by means of bridging aging cells and immune cells, and thus the cell adapter molecules of the present application can be used for removing aging cells expressing the NKG2D ligand and treating aging-related diseases.

The present invention relates to NK cells expressing dual-targeting chimeric antigen receptors for CD19 and CD22, and uses thereof. In the present invention, it was found that the delivery efficiency of a CAR expression vector and the production efficiency of CAR-NK cells are excellent when, among the cell surface proteins of PBMCs for producing CAR-NK cells (UCI-101), CD16 is expressed at 70% or less, natural killer group 2D (NKG2D) is expressed at less than 10%, CD57 is expressed at 30% or less, low-density lipoprotein receptor (LDLR) is expressed at 0.1% or more, and natural cytotoxicity triggering receptor 3 (NKp30) is expressed at less than 10%. In addition, optimal conditions for inducing NK cell differentiation and optimal conditions for transduction, under which the delivery efficiency of a CAR expression vector and the production efficiency of CAR-NK cells are excellent, were established, and CAR-NK cells produced by the method according to the present invention were found to exhibit an excellent antitumor effect in an animal model, and thus can be effectively used as a composition for preventing or treating diseases related to CD22 (or CD19) expression or diseases related to B cells.

The application belongs to the technical field of biological medicine, and particularly relates to application of a KAT5 inhibitor in a medicine for enhancing the immunotherapy effect of natural killer cells, and a medicine for enhancing the immunotherapy effect of natural killer cells. The KAT5 inhibitor comprises at least one of Nu9056, TH1834 and MG149. The KAT5 inhibitor can inhibit the recruitment of a transcription factor SP1 to an ADAM10 promoter, reduce the transcriptional activity of ADAM10, reduce the expression of NKG2D ligands on the cell surface, and thus enhance the sensitivity of tumor cells to NK cell killing. The application first discloses a new mechanism in which KAT5 regulates an ADAM10-NKG2D signal axis through an epigenetic mechanism, and proves that the KAT5 inhibitor can significantly enhance the immunotherapy effect of NK cells in vitro and in vivo. The application provides a brand-new target and strategy for developing a new immunosensitizer and a combined cell therapy.

The invention discloses an anti-NKG2D nano antibody, a fusion type nano antibody, a nucleic acid molecule, an expression vector, a recombinant cell and application thereof, and relates to the technical field of biological medicine. The anti-NKG2D nano antibody provided by the invention can specifically recognize and bind to NKG2D, can effectively competitively bind to a natural ligand (NKG2DL) of the NKG2D, and has relatively high affinity with the NKG2D. In-vitro experiments show that the provided antibody can effectively bind and activate NK cells so as to generate a remarkable killing effect on tumor cells. Therefore, the nano antibody provided by the invention can be applied to preparation of drugs for preventing or treating human tumors. In addition, the invention also provides a corresponding expression vector, a recombinant cell and an NKG2D detection product, an NKG2D enriched product and an NKG2D purified product related to the anti-NKG2D nano antibody, and the NKG2D detection product, the NKG2D enriched product and the NKG2D purified product have a wide application prospect.

The invention relates to the technical field of biological medicine, and particularly discloses a chimeric antigen receptor of a targeted NKG2D ligand, a nucleic acid molecule, an expression vector, a macrophage and a preparation method and application of the chimeric antigen receptor, the nucleic acid molecule, the expression vector and the macrophage, and an extracellular antigen binding domain is an NKG2D extracellular structural domain; the hinge region is a CD28 hinge region or a CD8a hinge region; the transmembrane / intracellular signal transduction structural domain is an FCGR1 transmembrane / intracellular signal transduction structural domain or a CD3z transmembrane / intracellular signal transduction structural domain. The chimeric antigen receptor provided by the invention specifically recognizes and is combined with NKG2DLs highly expressed in NKG2DL positive malignant tumors, and macrophages expressing the chimeric antigen receptor can directly swallow tumor cells, promote M1 type polarization, reprogram tumor microenvironment and induce recruitment and activation of T cells, so that effective treatment of the malignant tumors is realized.

Described herein are pharmaceutical formulations comprising multi-specific binding proteins that bind to NKG2D, CD16, and epidermal growth factor receptor (EGFR); and therapeutic uses of the multi-specific binding proteins and pharmaceutical formulations thereof for treating a disease, for example, cancer, in a patient in need thereof.

The application provides application of psoralen in activating NK cells. The psoralen provided by the application is a coumarin component extracted from traditional Chinese medicine psoralea; through in-vitro cell experiment verification, the psoralen can activate NK-92 to secrete IFN-gamma, Granzyme B and Perforion, and activate surface protein NKG2D of NK-92; meanwhile, the psoralen also has the same effect on a cell group containing NK cells; through in-vivo experiment verification, the psoralen helps to improve the total amount of NK cells in the mouse spleen, inhibit tumor growth, and improve the expression amount of IFN-gamma, Granzyme B, Perforion, NKG2D, NKP46 and CD107a in a tumor infiltration environment. In summary, the small-molecule compound psoralen has good effect in activating NK cells.

This invention belongs to the field of biotechnology, specifically relating to a high-affinity anti-tumor NK cell, its preparation method, and its application. This invention designs NK92 cells transfected with a haPD1 high-affinity chimeric conversion receptor, wherein the high-affinity chimeric conversion receptor includes the extracellular segment of haPD-1, the transmembrane segment of CD28, the intracellular segment of DAP10, and the intracellular segment of CD3ζ. This invention prepares haChR3-NK92 cells through the construction of a recombinant lentiviral vector, lentiviral packaging, and lentiviral transfection of NK92 cells. In this invention, haPD-1 serves as the extracellular recognition domain of the CAR structure, specifically binding to PD-L1 on tumor cells to achieve a stronger tumor-killing effect. The co-stimulatory molecule CD28 serves as the transmembrane region to transmit extracellular information into the cell, integrating the adaptor protein DAP10 of the NK cell surface activator receptor NKG2D into the cell, and further embedding the intracellular segment CD3ζ commonly used in CAR design to jointly promote NK cell activation. The preparation method of this invention can successfully construct haChR3-NK92 cells, achieve the expression of the target plasmid, and be applied in anti-tumor drug research.

The application discloses a sequential anti-tumor strategy based on improvement of tumor microenvironment (TME) combined with CAR T and application thereof, and belongs to the technical field of biological medicine, which simultaneously improves tumor fibrosis and acid microenvironment by applying tumor-related fibroblast (CAF) targeting drugs and proton pump inhibitors, and sequentially injects CAR T combined therapy for solid tumors, wherein the CAF targeting drugs are selected from tretinoin, calcipotriol, losartan and minnelide, the proton pump inhibitors are selected from lansoprazole, omeprazole, pantoprazole, rabeprazole and esomeprazole, and the CAR T receptor or ligand is selected from mesothelin, protein tyrosine kinase 7, NKG2D, CD47, B7-H3, MUC1 and HER2; the sequential administration strategy first destroys the dense physical barrier of the tumor microenvironment, improves the acidity of the tumor site, ensures the infiltration, survival and proliferation of CAR T at the tumor site, and then targets and inhibits the growth of solid tumors, especially high-malignancy triple-negative breast cancer, by using the specificity of CAR T.

The present application relates to an index combination, method and system for predicting obesity-related metabolic syndrome, which comprises an index combination for predicting obesity-related metabolic syndrome, the index combination comprising the following indexes: NKG2C + Percentage of mitochondrial low membrane potential in NK cells (NKG2C + NK.MMP low Percentage), NKG2D + Percentage of mitochondrial low membrane potential in NKT cells (NKG2D + NKT.MMP low Percentage), NKG2C + Percentage of mitochondrial low membrane potential in NKT cells (NKG2C + NKT.MMP low Percentage), percentage of mitochondrial low membrane potential in monocytes (Mono.MMP low Percentage), NKG2A + Mitochondrial mass in lymphocytes (NKG2A + Lymph.MM), and mean platelet volume (MPV). The index combination obtained by screening has high biological interpretability and clinical relevance, and can effectively reflect obesity-related immune metabolic abnormalities.

The invention relates to an index combination, a method and a system for predicting obesity-related metabolic syndrome, which comprise an index combination for predicting obesity-related metabolic syndrome. The index combination comprises the following indexes: the percentage of mitochondrial low membrane potential in NKG2C + NK cells (NKG2C + NK.MMPlow%), the percentage of mitochondrial low membrane potential in NKG2D + NKT cells (NKG2D + NKT.MMPlow%), the percentage of mitochondrial low membrane potential in NKG2C + NKT cells (NKG2C + NKT.MMPlow%), the percentage of mitochondrial low membrane potential in mononuclear cells (Mono.MMPlow%), the percentage of mitochondrial mass in NKG2A + lymphocyte (NKG2A + Lymp.MM) and the average platelet volume (MPV). The index combination obtained by screening has relatively high biological interpretation and clinical correlation, and can effectively reflect obesity-related immunometabolism abnormality.

This invention provides a method for screening peptides that specifically target T cells, comprising providing T cells having a subset selected from CD1, CD2, CD3, CD4, CD5, CD7, CD8, CD16, CD25, CD26, CD27, CD28, CD30, CD38, CD39, CD40L, CD44, CD45, CD62L, CD69, CD73, CD80, CD83, CD86, CD95, CD103, CD119, CD126, CD150, CD152 (CTLA-4), CD153, CD154 (CD40L) The T cells are labeled with at least one of the following surface antigen markers: CD161, CD183, CD223, CD254, CD275, CD45RA, CXCR3, CXCR5, FasL, IL18R1, CTLA-4, OX40, GITR, LAG3, ICOS, PD-1, leu-12, TCR, TLR1, TLR2, TLR3, TLR4, TLR6, NKG2D, CCR, CCR1, CCR2, CCR4, CCR6, and CCR7. The T cells are then contacted with a peptide display library, and target peptides that specifically bind to the T cells are screened therefrom.

The invention provides an application of in-vitro amplified NK cells in treatment of autoimmune diseases, the NK cells are obtained by performing in-vitro induced amplification on peripheral blood mononuclear cells, and the in-vitro induced amplification is performed in a culture medium containing interleukin and / or an antibody. Therefore, the NK cell disclosed by the invention has double advantages that on one hand, the NK cell shows phenotypic characteristics of a CD56light tissue resident type NK cell, has the capability of penetrating pathological tissues, and can be used for directly removing abnormal B cells and T cells in tissues of patients with autoimmune diseases; on the other hand, activated receptors such as NKG2D and CD16 and chemokine receptors such as CXCR3 can be stably and highly expressed, and the removal efficiency is further improved by enhancing the recognition and killing capacity on target cells.

The present invention relates to a pharmaceutical composition for preventing or treating cancer, the pharmaceutical composition containing CD4+ CTL as an active ingredient. The CD4+ CTL according to the present invention performs a cytotoxic function of directly attacking and killing myeloma cells, thereby extending the progression-free survival of transplant-ineligible myeloma patients, and thus can contribute to increasing the survival rate of the patients. In addition, by using NKG2D+ CD4+ CTL as a biomarker for predicting the prognosis of myeloma patients, it is possible to analyze the CD4+ CTL status of individual patients and thereby establish customized treatment strategies. In addition, by increasing the cytotoxicity of NKG2D+ CD4+ CTL through a specific cytokine treatment, it is possible to more effectively prevent or treat cancer.

Provided is a method for screening polypeptides that specifically target T cells, comprising: providing T cells having a surface antigen marker selected from at least one of CD1, CD2, CD3, CD4, CD5, CD7, CD8, CD16, CD25, CD26, CD27, CD28, CD30, CD38, CD39, CD40L, CD44, CD45, CD62L, CD69, CD73, CD80, CD83, CD86, CD95, CD103, CD119, CD126, CD150, CD152 (CTLA-4), CD153, CD154 (CD40L), CD161, CD183, CD223, CD254, CD275, CD45RA, CXCR3, CXCR5, FasL, IL18R1, CTLA-4, OX40, GITR, LAG3, ICOS, PD-1, leu-12, TCR, TLR1, TLR2, TLR3, TLR4, TLR6, NKG2D, CCR, CCR1, CCR2, CCR4, CCR6, and CCR7; and contacting the T cells with a polypeptide display library, and screening therefrom target polypeptides that specifically bind to the T cells.

The application discloses application of combination of CAR-T and CAR-M in preparation of antitumor drugs, wherein the CAR-T is a T cell modified by a chimeric antigen receptor, and the primary protein structure of the chimeric antigen receptor sequentially comprises a signal peptide, an NKG2D extracellular region, a hinge region, a transmembrane region, an intracellular costimulatory signal region and an intracellular signal region from an amino terminus to a carboxyl terminus; the CAR-M is a macrophage modified by a chimeric antigen receptor, and the primary protein structure of the chimeric antigen receptor sequentially comprises a signal peptide, an NKG2D extracellular region, a hinge region, a transmembrane region and an intracellular signal region from an amino terminus to a carboxyl terminus; and the NKG2D extracellular region is a human NKG2D extracellular domain, and the amino acid sequence is shown as SEQ ID NO. 2. The combination of the CAR-T and the CAR-M combines the ability of the CAR-M to improve a tumor microenvironment with the ability of the CAR-T to directly kill tumor cells, so that strong and efficient killing of tumor cells can be realized, and the curative effect is greatly improved.

A dual-targeting chimeric antigen receptor (CAR) and its encoding gene, a recombinant expression vector, natural killer cells, and applications thereof are provided, relating to the field of tumor immunotherapy. The dual-targeting CAR includes a dual-CAR molecular encoding sequence sequentially connected encoding sequences of: an extracellular antigen-binding region, a transmembrane region, and an intracellular signal transduction region. PD1 molecule extracellular domain and NKG2D molecule extracellular domain serve as a first recognition target and a second recognition target respectively. When the PD1 molecule extracellular domain recognizes PD-L1 molecules on tumor cells, the negative regulatory signal is transmitted downward, triggering activation of the DAP10 activating receptor pathway and converting it into a positive activation signal. Simultaneously, when the NKG2D molecule extracellular domain recognizes NKG2DL molecules on tumor cells, NK cell activation is induced. Under the synergistic action of costimulatory molecules 4-1BB and CD3ζ, the anti-tumor therapeutic efficacy of NK cells is significantly enhanced.

The invention belongs to the technical field of biology, and particularly relates to a novel chimeric antigen receptor modified immune cell and application of the novel chimeric antigen receptor modified immune cell in preparation of drugs, and the drugs can improve the anti-tumor cell factor secretion capacity of the immune cell; wherein the immune cell is modified with a chimeric antigen receptor, and the chimeric antigen receptor comprises an extracellular region, a transmembrane region and an intracellular region; the extracellular region can be combined with a tumor antigen; the transmembrane region is connected with the extracellular region; the intracellular region is connected with the transmembrane region; the intracellular region is selected from all amino acid sequences or partial amino acid sequences of at least one of the following proteins: 2B4, DAP10, CD3 zeta and SAP; the extracellular region is derived from an NK cell activation receptor; the tumor antigen is a tumor specific antigen or a tumor associated antigen. According to the application, the novel chimeric antigen receptor modified immune cell is applied to preparation of anti-tumor drugs, and by means of the innovatively designed chimeric antigen receptor, the anti-tumor cell factor secretion capacity of the immune cell is remarkably enhanced; according to the present invention, the CAR-immune cell immunotherapy kit has characteristics of high sensitivity and high efficiency, can accurately identify and efficiently attack a variety of NKG2D ligand positive tumor cells, can reduce the non-specific attack on normal cells, can significantly improve the safety and the effectiveness when the CAR-immune cell immunotherapy kit is used for treatment, and can provide a new idea for the CAR-immune cell therapy.

This invention discloses a culture medium that can significantly improve the in vitro expansion efficiency and anti-tumor activity of NK cells. The culture medium comprises: (1) a basal medium: RPMI 1640 medium containing 10% human AB serum, 1% GlutaMAX, and 0.1% β-mercaptoethanol; (2) a fusion polypeptide: with a final concentration of 150 ng / mL, which achieves synergistic activation of the IL-15R and NKG2D dual pathways by linking the IL-15 mutant with the NKG2D ligand ULBP2; and (3) an anti-PD-1 antibody: with a final concentration of 30 μg / mL, which blocks the PD-1 / PD-L1 inhibitory signal. Experiments show that after 7 days of culture in this medium, NK cells can expand by 39.8 times, the CD69 activation rate increases to 83.6%, and the apoptosis rate decreases to 6.5%. The in vitro killing rate of PD-L1+ tumor cells is increased to 64.7%, the tumor volume inhibition rate in tumor-bearing mouse models reaches 66.4%, and the median survival time is extended to >60 days. This invention overcomes the limitations of existing NK cell expansion and activity through multi-pathway synergy, providing a highly efficient solution for immunotherapy of solid tumors.

The invention relates to a method for evaluating the immune state of obese people and a kit thereof, relates to the technical field of medical data mining, and comprises an index combination for evaluating the immune state of the obese people. The index combination comprises the following indexes: hematocrit (HCT), hemoglobin content (HGB), low membrane potential percentage (Ts.MMPlow%) of killer T cells, percentage (T4Tcm%) of central memory assistant T cells, low membrane potential percentage (T8Tn.MMPlow%) of pure killer T cell mitochondria, percentage (NKT.NKG2D +%) of NKT cell expression NKG2D and mitochondrial mass (T8Tcm. MM) of central memory killer T cell mitochondria. The index combination obtained through screening has high biological interpretation and clinical correlation, and the immune state of obese people can be effectively evaluated.

The application discloses a multifunctional immunoregulatory fusion protein, a preparation method thereof, NK cells expressing the protein and application of the protein. The fusion protein structure is anti-CD16a VHH-(GGGGS)3-anti-NKG2D VHH-(EAAAK)2-IL-15R beta super agonist peptide, and the amino acid sequence is SEQ ID NO:1. The anti-CD16a VHH and the anti-NKG2D VHH synergistically activate immune effector cells, the IL-15R beta super agonist peptide strongly activates effector cells and weakly activates Tregs, and the linker ensures that the functions of the domains are independent. The protein can be efficiently prepared, the yield is 155 mg / L, the purity is higher than 95%, the NK cells and CD8 + T cells can be significantly activated, the 24h lysis rate of the protein on various tumor cells reaches 87.0%-90.0% in vitro, the tumor growth is significantly inhibited in vivo, the inhibition rate is 81.3%, and the survival period of tumor-bearing mice is prolonged by more than 60 days. The stability of the NK cells expressing the protein is improved after being treated by MI-3, and the protein provides a high-efficiency and safe candidate drug for tumor immunotherapy.

The present invention provides combination therapy using gamma delta (γδ) T cells (e.g., Vδ1+) for treating solid cancers. In particular, the present invention provides several strategies to enhance efficacy of gamma delta (γδ) T cells against cancer cells. The synergistical strategies include combination therapy of gamma delta (γδ) T cell therapy and immune checkpoint blockade immunotherapy and / or upregulation of NKG2D reaction to increase gamma delta (γδ) T cells targeting cancer tissues.

The present disclosure relates to the treatment of cancer. In particular, the present disclosure provides novel therapeutic antibodies that target MHC class I chain-associated protein A (MICA) in a manner that reduces the ability of tumor cells to escape immune clearance mediated by natural killer cell group 2D (NKG2D) receptors.

This invention belongs to the field of antibody preparation technology, specifically relating to an anti-NKG2D nanobody, its preparation method, and its application. The method is as follows: NKG2D protein expression is constructed, and alpacas are immunized to obtain immunized alpaca PBMC cells. RNA is extracted from the PBMC cells, reverse transcribed into cDNA, and antibody gene fragments are obtained by PCR. These fragments are then constructed into a phage expression vector. The obtained vector containing the antibody gene fragment is electroporated into SS320 electrocompetent cells to obtain an NKG2D immune antibody library. After two rounds of NKG2D protein panning and Clone ELISA detection, antibodies binding to NKG2D protein are obtained. After sequencing analysis, unique sequences are selected and expressed in a mammalian cell high-throughput expression system to obtain antibody proteins. The antibodies are then detected using an NKG2D high-expression cell line via FACS, ultimately yielding antibodies binding to NKG2D. The method of this invention is simple and rapid; the mammalian cell expression system induces highly efficient antibody expression, and the translated antibodies can be further modified, resulting in activity closer to that of natural antibodies.

The application provides an application of dodartin A1 in activating NK cells. Through in-vitro experiments, it is proved that dodartin A1 can significantly promote the synthesis and release of IFN-gamma and cytotoxic substances (such as granzyme B and perforin) of NK92 cells, and increase the expression of the surface agonistic receptors NKp46 and NKG2D of the NK92 cells; through in-vivo experiments, it is proved that dodartin A1 can promote the expression of the NK cell activation related substances CD107a and NKp46 to play an anti-hepatoma effect; in conclusion, dodartin A1 has the effect of activating NK cells.

The present invention relates to a fusion protein comprising (a) an antibody or antibody fragment that binds to an antigen expressed on the surface of a target cell of an NK cell, a T cell and / or an NKT cell, preferably a B cell or a tumor cell, (b) an antibody or antibody fragment that binds to 4-1BB, NKG2D, NKp30, NKp46, NKp44, 2B4, CD28 or DNAM1, and (c) IL-15, IL-2, IL-18, IL-21 or IL-12.

The invention relates to an enhanced chimeric antigen receptor-T cell and a preparation method and application thereof.A nucleotide sequence for coding a chimeric antigen receptor is exogenously inserted into a genome of the T cell, and the nucleotide sequence comprises at least one of the following genes: an NKG2D gene, a DNAM1 gene, an NKp30 gene, an NKp44 gene or an NKp46 gene, therefore, the expression level of the NKG2D protein, the DNAM1 protein, the NKp30 protein, the NKp44 protein or the NKp46 protein of the chimeric antigen receptor-T cell is higher than the expression level of the corresponding protein in the original T cell. The enhanced chimeric antigen receptor-T cell can more effectively attack tumor cells in a targeted manner, has high tumor killing rate, does not depend on specific tumor antigens and specific tumor mutation, is expected to break through antigen escape, and can be used for preparing anti-tumor products.