CN113189340A - 细胞焦亡通路在细胞治疗中的用途 - Google Patents
细胞焦亡通路在细胞治疗中的用途 Download PDFInfo
- Publication number
- CN113189340A CN113189340A CN202010035350.7A CN202010035350A CN113189340A CN 113189340 A CN113189340 A CN 113189340A CN 202010035350 A CN202010035350 A CN 202010035350A CN 113189340 A CN113189340 A CN 113189340A
- Authority
- CN
- China
- Prior art keywords
- gsdme
- cell
- protein
- lymphoma
- agent
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 230000006907 apoptotic process Effects 0.000 title abstract description 24
- 230000037361 pathway Effects 0.000 title abstract description 6
- 238000002659 cell therapy Methods 0.000 title abstract description 4
- 101001026269 Homo sapiens Gasdermin-E Proteins 0.000 claims abstract description 90
- 102100037391 Gasdermin-E Human genes 0.000 claims abstract description 81
- 206010052015 cytokine release syndrome Diseases 0.000 claims abstract description 55
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 41
- 230000000694 effects Effects 0.000 claims abstract description 40
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 34
- 230000002401 inhibitory effect Effects 0.000 claims abstract description 7
- 238000004519 manufacturing process Methods 0.000 claims abstract description 7
- 230000001603 reducing effect Effects 0.000 claims abstract description 6
- 239000003814 drug Substances 0.000 claims abstract description 4
- 206010028980 Neoplasm Diseases 0.000 claims description 32
- 201000011510 cancer Diseases 0.000 claims description 27
- 210000001744 T-lymphocyte Anatomy 0.000 claims description 23
- 230000014509 gene expression Effects 0.000 claims description 23
- 102100024222 B-lymphocyte antigen CD19 Human genes 0.000 claims description 22
- 101000980825 Homo sapiens B-lymphocyte antigen CD19 Proteins 0.000 claims description 22
- 101001012157 Homo sapiens Receptor tyrosine-protein kinase erbB-2 Proteins 0.000 claims description 22
- 102100030086 Receptor tyrosine-protein kinase erbB-2 Human genes 0.000 claims description 22
- 208000025324 B-cell acute lymphoblastic leukemia Diseases 0.000 claims description 20
- 238000011357 CAR T-cell therapy Methods 0.000 claims description 16
- 208000017815 Dendritic cell tumor Diseases 0.000 claims description 14
- 201000003444 follicular lymphoma Diseases 0.000 claims description 14
- 108020004999 messenger RNA Proteins 0.000 claims description 13
- 208000025205 Mantle-Cell Lymphoma Diseases 0.000 claims description 11
- 208000009052 Precursor T-Cell Lymphoblastic Leukemia-Lymphoma Diseases 0.000 claims description 11
- 208000029052 T-cell acute lymphoblastic leukemia Diseases 0.000 claims description 11
- 201000003793 Myelodysplastic syndrome Diseases 0.000 claims description 10
- 210000003719 b-lymphocyte Anatomy 0.000 claims description 10
- 210000005134 plasmacytoid dendritic cell Anatomy 0.000 claims description 10
- 230000003211 malignant effect Effects 0.000 claims description 8
- 208000036762 Acute promyelocytic leukaemia Diseases 0.000 claims description 7
- 208000036170 B-Cell Marginal Zone Lymphoma Diseases 0.000 claims description 7
- 208000010839 B-cell chronic lymphocytic leukemia Diseases 0.000 claims description 7
- 208000032791 BCR-ABL1 positive chronic myelogenous leukemia Diseases 0.000 claims description 7
- 208000011691 Burkitt lymphomas Diseases 0.000 claims description 7
- 206010061850 Extranodal marginal zone B-cell lymphoma (MALT type) Diseases 0.000 claims description 7
- 208000031671 Large B-Cell Diffuse Lymphoma Diseases 0.000 claims description 7
- 206010025280 Lymphocytosis Diseases 0.000 claims description 7
- 201000003791 MALT lymphoma Diseases 0.000 claims description 7
- 208000034578 Multiple myelomas Diseases 0.000 claims description 7
- 208000015914 Non-Hodgkin lymphomas Diseases 0.000 claims description 7
- 206010035226 Plasma cell myeloma Diseases 0.000 claims description 7
- 208000033559 Waldenström macroglobulinemia Diseases 0.000 claims description 7
- 206010012818 diffuse large B-cell lymphoma Diseases 0.000 claims description 7
- 201000009277 hairy cell leukemia Diseases 0.000 claims description 7
- 201000007924 marginal zone B-cell lymphoma Diseases 0.000 claims description 7
- 208000021937 marginal zone lymphoma Diseases 0.000 claims description 7
- 239000000203 mixture Substances 0.000 claims description 7
- 108090000397 Caspase 3 Proteins 0.000 claims description 6
- 102100029855 Caspase-3 Human genes 0.000 claims description 6
- 239000003153 chemical reaction reagent Substances 0.000 claims description 6
- 102100035904 Caspase-1 Human genes 0.000 claims description 5
- 108090000426 Caspase-1 Proteins 0.000 claims description 5
- 101710090338 Caspase-4 Proteins 0.000 claims description 5
- 102100025597 Caspase-4 Human genes 0.000 claims description 5
- 102100038916 Caspase-5 Human genes 0.000 claims description 5
- 101710090333 Caspase-5 Proteins 0.000 claims description 5
- 101000933115 Mus musculus Caspase-4 Proteins 0.000 claims description 5
- 230000000692 anti-sense effect Effects 0.000 claims description 4
- 239000003112 inhibitor Substances 0.000 claims description 4
- 229940076372 protein antagonist Drugs 0.000 claims description 3
- 208000010833 Chronic myeloid leukaemia Diseases 0.000 claims 3
- 208000031422 Lymphocytic Chronic B-Cell Leukemia Diseases 0.000 claims 3
- 208000033761 Myelogenous Chronic BCR-ABL Positive Leukemia Diseases 0.000 claims 3
- 208000007452 Plasmacytoma Diseases 0.000 claims 3
- 208000032852 chronic lymphocytic leukemia Diseases 0.000 claims 3
- 230000004952 protein activity Effects 0.000 claims 2
- 101100314454 Caenorhabditis elegans tra-1 gene Proteins 0.000 claims 1
- 230000034994 death Effects 0.000 abstract description 10
- 230000001404 mediated effect Effects 0.000 abstract description 8
- 210000004027 cell Anatomy 0.000 description 69
- 108091007741 Chimeric antigen receptor T cells Proteins 0.000 description 15
- 210000004881 tumor cell Anatomy 0.000 description 15
- 102000004121 Annexin A5 Human genes 0.000 description 11
- 108090000672 Annexin A5 Proteins 0.000 description 11
- 208000032839 leukemia Diseases 0.000 description 10
- 210000003462 vein Anatomy 0.000 description 10
- 208000024893 Acute lymphoblastic leukemia Diseases 0.000 description 9
- 208000014697 Acute lymphocytic leukaemia Diseases 0.000 description 9
- 208000006664 Precursor Cell Lymphoblastic Leukemia-Lymphoma Diseases 0.000 description 9
- 241000699670 Mus sp. Species 0.000 description 8
- 230000002757 inflammatory effect Effects 0.000 description 8
- 210000002966 serum Anatomy 0.000 description 8
- 210000002540 macrophage Anatomy 0.000 description 7
- 208000024891 symptom Diseases 0.000 description 7
- 102000017420 CD3 protein, epsilon/gamma/delta subunit Human genes 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 230000004083 survival effect Effects 0.000 description 6
- 208000004736 B-Cell Leukemia Diseases 0.000 description 5
- 238000008157 ELISA kit Methods 0.000 description 5
- 101150071723 GSDME gene Proteins 0.000 description 5
- 101001011663 Homo sapiens Mixed lineage kinase domain-like protein Proteins 0.000 description 5
- 108090001005 Interleukin-6 Proteins 0.000 description 5
- 102100030177 Mixed lineage kinase domain-like protein Human genes 0.000 description 5
- 241000699666 Mus <mouse, genus> Species 0.000 description 5
- 238000000684 flow cytometry Methods 0.000 description 5
- 238000002347 injection Methods 0.000 description 5
- 239000007924 injection Substances 0.000 description 5
- 230000017074 necrotic cell death Effects 0.000 description 5
- 239000006228 supernatant Substances 0.000 description 5
- 108010083359 Antigen Receptors Proteins 0.000 description 4
- 102000006306 Antigen Receptors Human genes 0.000 description 4
- 101000914514 Homo sapiens T-cell-specific surface glycoprotein CD28 Proteins 0.000 description 4
- 102000003777 Interleukin-1 beta Human genes 0.000 description 4
- 108090000193 Interleukin-1 beta Proteins 0.000 description 4
- 101710190759 Serum amyloid A protein Proteins 0.000 description 4
- 238000000692 Student's t-test Methods 0.000 description 4
- 102100027213 T-cell-specific surface glycoprotein CD28 Human genes 0.000 description 4
- 238000001415 gene therapy Methods 0.000 description 4
- 238000000338 in vitro Methods 0.000 description 4
- 108010027775 interleukin-1beta-converting enzyme inhibitor Proteins 0.000 description 4
- 238000001543 one-way ANOVA Methods 0.000 description 4
- 239000013612 plasmid Substances 0.000 description 4
- 239000002516 radical scavenger Substances 0.000 description 4
- 108010019670 Chimeric Antigen Receptors Proteins 0.000 description 3
- 102000004127 Cytokines Human genes 0.000 description 3
- 108090000695 Cytokines Proteins 0.000 description 3
- 102100037388 Gasdermin-D Human genes 0.000 description 3
- 101100383038 Homo sapiens CD19 gene Proteins 0.000 description 3
- 101001026262 Homo sapiens Gasdermin-D Proteins 0.000 description 3
- 101001109145 Homo sapiens Receptor-interacting serine/threonine-protein kinase 1 Proteins 0.000 description 3
- 102100022501 Receptor-interacting serine/threonine-protein kinase 1 Human genes 0.000 description 3
- 210000000170 cell membrane Anatomy 0.000 description 3
- 238000010276 construction Methods 0.000 description 3
- 230000002147 killing effect Effects 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 238000004806 packaging method and process Methods 0.000 description 3
- 102000005962 receptors Human genes 0.000 description 3
- 108020003175 receptors Proteins 0.000 description 3
- 238000002560 therapeutic procedure Methods 0.000 description 3
- 231100000155 toxicity by organ Toxicity 0.000 description 3
- 230000007675 toxicity by organ Effects 0.000 description 3
- 238000010361 transduction Methods 0.000 description 3
- 230000026683 transduction Effects 0.000 description 3
- 241001430294 unidentified retrovirus Species 0.000 description 3
- 208000003950 B-cell lymphoma Diseases 0.000 description 2
- SJDDOCKBXFJEJB-MOKWFATOSA-N Belnacasan Chemical compound CCO[C@@H]1OC(=O)C[C@@H]1NC(=O)[C@H]1N(C(=O)[C@@H](NC(=O)C=2C=C(Cl)C(N)=CC=2)C(C)(C)C)CCC1 SJDDOCKBXFJEJB-MOKWFATOSA-N 0.000 description 2
- 238000010356 CRISPR-Cas9 genome editing Methods 0.000 description 2
- 229940124101 Caspase 3 inhibitor Drugs 0.000 description 2
- 108020004414 DNA Proteins 0.000 description 2
- 102000017727 Immunoglobulin Variable Region Human genes 0.000 description 2
- 108010067060 Immunoglobulin Variable Region Proteins 0.000 description 2
- 238000012404 In vitro experiment Methods 0.000 description 2
- 241000713666 Lentivirus Species 0.000 description 2
- 241001529936 Murinae Species 0.000 description 2
- 206010037660 Pyrexia Diseases 0.000 description 2
- 230000004913 activation Effects 0.000 description 2
- 238000003782 apoptosis assay Methods 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
- 239000011324 bead Substances 0.000 description 2
- 229950010527 belnacasan Drugs 0.000 description 2
- 230000000903 blocking effect Effects 0.000 description 2
- 230000001413 cellular effect Effects 0.000 description 2
- 229940044683 chemotherapy drug Drugs 0.000 description 2
- 230000002596 correlated effect Effects 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 238000001802 infusion Methods 0.000 description 2
- 239000006166 lysate Substances 0.000 description 2
- 230000036210 malignancy Effects 0.000 description 2
- 108091070501 miRNA Proteins 0.000 description 2
- 239000002679 microRNA Substances 0.000 description 2
- 230000009456 molecular mechanism Effects 0.000 description 2
- 230000036457 multidrug resistance Effects 0.000 description 2
- 229910052760 oxygen Inorganic materials 0.000 description 2
- 239000001301 oxygen Substances 0.000 description 2
- 239000011148 porous material Substances 0.000 description 2
- 230000005522 programmed cell death Effects 0.000 description 2
- 230000000770 proinflammatory effect Effects 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108010056030 retronectin Proteins 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000001262 western blot Methods 0.000 description 2
- 108091033409 CRISPR Proteins 0.000 description 1
- 108010076667 Caspases Proteins 0.000 description 1
- 102000011727 Caspases Human genes 0.000 description 1
- 206010050685 Cytokine storm Diseases 0.000 description 1
- 102100037387 Gasdermin-A Human genes 0.000 description 1
- 206010019233 Headaches Diseases 0.000 description 1
- 101000643956 Homo sapiens Cytochrome b-c1 complex subunit Rieske, mitochondrial Proteins 0.000 description 1
- 101001026276 Homo sapiens Gasdermin-A Proteins 0.000 description 1
- 101001002657 Homo sapiens Interleukin-2 Proteins 0.000 description 1
- 101001099199 Homo sapiens RalA-binding protein 1 Proteins 0.000 description 1
- 101000964436 Homo sapiens Z-DNA-binding protein 1 Proteins 0.000 description 1
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 1
- 208000001953 Hypotension Diseases 0.000 description 1
- 102100037924 Insulin-like growth factor 2 mRNA-binding protein 1 Human genes 0.000 description 1
- 108010002352 Interleukin-1 Proteins 0.000 description 1
- 102000000589 Interleukin-1 Human genes 0.000 description 1
- 108010002350 Interleukin-2 Proteins 0.000 description 1
- 241000581650 Ivesia Species 0.000 description 1
- 206010024380 Leukoderma Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000525154 Micrathena beta Species 0.000 description 1
- 102000014842 Multidrug resistance proteins Human genes 0.000 description 1
- 108050005144 Multidrug resistance proteins Proteins 0.000 description 1
- 101000858032 Mus musculus Coxsackievirus and adenovirus receptor homolog Proteins 0.000 description 1
- 101001124276 Mus musculus Nuclear receptor subfamily 1 group I member 3 Proteins 0.000 description 1
- 208000000112 Myalgia Diseases 0.000 description 1
- 101710156256 Myosin phosphatase Rho-interacting protein Proteins 0.000 description 1
- 108091008638 NR4A Proteins 0.000 description 1
- 206010028813 Nausea Diseases 0.000 description 1
- 239000000020 Nitrocellulose Substances 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 102000001253 Protein Kinase Human genes 0.000 description 1
- 101710092490 Protein kinase 3 Proteins 0.000 description 1
- 101000599776 Rattus norvegicus Insulin-like growth factor 2 mRNA-binding protein 1 Proteins 0.000 description 1
- 102100033729 Receptor-interacting serine/threonine-protein kinase 3 Human genes 0.000 description 1
- 108091027544 Subgenomic mRNA Proteins 0.000 description 1
- 230000006044 T cell activation Effects 0.000 description 1
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 1
- 102100040247 Tumor necrosis factor Human genes 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 230000010398 acute inflammatory response Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 239000005557 antagonist Substances 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 238000003339 best practice Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000003915 cell function Effects 0.000 description 1
- 208000015114 central nervous system disease Diseases 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000000875 corresponding effect Effects 0.000 description 1
- 230000000139 costimulatory effect Effects 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 239000012636 effector Substances 0.000 description 1
- 206010016256 fatigue Diseases 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000003209 gene knockout Methods 0.000 description 1
- 231100000869 headache Toxicity 0.000 description 1
- 210000002216 heart Anatomy 0.000 description 1
- 230000004217 heart function Effects 0.000 description 1
- 238000011577 humanized mouse model Methods 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000036543 hypotension Effects 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 230000004068 intracellular signaling Effects 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 230000003907 kidney function Effects 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 230000002101 lytic effect Effects 0.000 description 1
- 206010025482 malaise Diseases 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000003990 molecular pathway Effects 0.000 description 1
- 238000010172 mouse model Methods 0.000 description 1
- 230000008693 nausea Effects 0.000 description 1
- 230000001338 necrotic effect Effects 0.000 description 1
- 210000003061 neural cell Anatomy 0.000 description 1
- 229920001220 nitrocellulos Polymers 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- -1 polyethylene Polymers 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000004393 prognosis Methods 0.000 description 1
- 108060006633 protein kinase Proteins 0.000 description 1
- 230000007115 recruitment Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 230000000241 respiratory effect Effects 0.000 description 1
- 230000001177 retroviral effect Effects 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- 230000008718 systemic inflammatory response Effects 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 230000001960 triggered effect Effects 0.000 description 1
- 210000003934 vacuole Anatomy 0.000 description 1
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4611—T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/463—Cellular immunotherapy characterised by recombinant expression
- A61K39/4631—Chimeric Antigen Receptors [CAR]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/464—Cellular immunotherapy characterised by the antigen targeted or presented
- A61K39/4643—Vertebrate antigens
- A61K39/4644—Cancer antigens
- A61K39/464402—Receptors, cell surface antigens or cell surface determinants
- A61K39/464403—Receptors for growth factors
- A61K39/464406—Her-2/neu/ErbB2, Her-3/ErbB3 or Her 4/ ErbB4
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/464—Cellular immunotherapy characterised by the antigen targeted or presented
- A61K39/4643—Vertebrate antigens
- A61K39/4644—Cancer antigens
- A61K39/464402—Receptors, cell surface antigens or cell surface determinants
- A61K39/464411—Immunoglobulin superfamily
- A61K39/464412—CD19 or B4
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
- A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
- A61P35/02—Antineoplastic agents specific for leukemia
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
- C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/31—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterized by the route of administration
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/38—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterised by the dose, timing or administration schedule
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/46—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterised by the cancer treated
- A61K2239/48—Blood cells, e.g. leukemia or lymphoma
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/158—Expression markers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/56—Staging of a disease; Further complications associated with the disease
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Immunology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Microbiology (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Cell Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Mycology (AREA)
- Oncology (AREA)
- Analytical Chemistry (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Pathology (AREA)
- General Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Urology & Nephrology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Biomedical Technology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biophysics (AREA)
- Hospice & Palliative Care (AREA)
- Food Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Physics & Mathematics (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
本发明涉及细胞焦亡通路在细胞治疗中的用途。具体地，本发明涉及Gasdermin E介导的焦亡通路在细胞因子释放综合征的预测和/或治疗中的用途。具体地，本发明涉及特异性检测GSDME蛋白质或基因活性或水平的试剂在制备用于预测受试者发生细胞因子释放综合征的风险的试剂盒中的用途，另一方面，本发明涉及阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂在制备用于抑制和/或降低受试者发生细胞因子释放综合征的药物中的用途。
Description
技术领域
本发明涉及细胞因子释放综合征预测和/或治疗的技术领域。具体地，本发明涉及Gasdermin E(GSDME)作为标记物预测细胞因子释放综合征的用途，以及阻断或抑制Gasdermin E(GSDME)基因表达和/或激活通路来改善细胞因子释放综合征的技术领域。
背景技术
运用嵌合抗原受体修饰的基因工程T细胞(CAR-T)在治疗恶性肿瘤，例如B细胞恶性肿瘤的临床治疗取得了显著疗效(参见S.S.Neelapu等人，Axicabtagene CiloleucelCAR T-Cell Therapy in Refractory Large B-Cell Lymphoma.N Engl J Med 377,2531-2544(2017)；S.L.Maude等人，Tisagenlecleucel in Children and Young Adults withB-Cell Lymphoblastic Leukemia.N Engl J 17Med 378,439-448(2018)；和M.Sadelain等人,Therapeutic T cell engineering.Nature 545,19 423-431(2017))，但治疗过程中往往伴发细胞因子释放综合征(CRS)。这种严重的全身性炎症反应阻碍了CAR-T细胞临床治疗的进一步发展(参见，C.L.Bonifant等人Toxicity and management in CAR T-celltherapy.Mol Ther Oncolytics 3,16011(2016)；M.L.Davila,等人Efficacy andtoxicity management of 19-28z CAR T cell therapy in B cell acutelymphoblastic leukemia.Sci Transl Med 6,224ra225(2014))。目前研究表明，CRS是一种急性炎症反应，其特征表现为血清炎性细胞因子升高及相关的发烧，低血压和呼吸功能不全等临床症状(参见N.Frey,Cytokine release syndrome:Who is at risk and how totreat.Best Pract 8Res Clin Haematol 30,336-340(2017)；D.T.Teachey,等人Identification of Predictive Biomarkers for Cytokine Release Syndrome afterChimeric Antigen Receptor T-cell Therapy for Acute LymphoblasticLeukemia.Cancer Discov 6,664-679(2016)；和M.L.Davila等人Efficacy and toxicitymanagement of 19-28z CAR T cell therapy in B cell acute lymphoblasticleukemia.Sci Transl Med 6,224ra225-224ra225(2014))。CAR-T细胞在输注病人体内后快速激活扩增，扩增后的CAR-T细胞在短时间内导致B白血病细胞的迅速大量死亡。CRS的发生率及症状严重程度与B白血病细胞的裂解破坏程度成正相关。但这一急性恶性肿瘤细胞大量坏死与CRS发生的内在关联的分子机制尚未探究清楚。同时，另有研究表明，在CAR-T细胞治疗的人源化小鼠模型中，巨噬细胞参与了CRS的发生(参见T.Giavridis等人，CAR Tcell-induced cytokine release syndrome is mediated by macrophages and abatedby IL-1 blockade.Nat Med 24,731-738(2018)；和M.Norelli等人Monocyte-derived IL-1and IL-6are differentially required for cytokine-release syndrome andneurotoxicity due to CAR T cells.Nat Med 24,739-748(2018))，但其具体分子机制仍不清楚。
细胞有多种死亡方式。凋亡一直被认为是细胞程序性死亡的唯一形式。但目前的研究表明了新形式的程序性死亡的存在，其特征是细胞迅速肿胀，质膜上出现大的胞膜泡以及促炎因子的释放(参见J.Shi,等人,Pyroptosis:Gasdermin-Mediated ProgrammedNecrotic Cell Death.Trends Biochem Sci 42,245-254(2017)；和D.Wallach等人Programmed necrosis in inflammation:Toward identification of the effectormolecules.Science 352,aaf2154(2016))。到目前为止，至少已经确定了两个新的介导程序性细胞死亡的分子途径。一种是混合谱系激酶结构域样假激酶(MLKL)介导的坏死性死亡(参见J.Lin等人RIPK1 counteracts ZBP1-mediated necroptosis to inhibitinflammation.Nature 540,124-128(2016)；和J.Yuan,等人Necroptosis and RIPK1-mediated neuroinflammation in CNS diseases.Nat Rev Neurosci 20,19-33(2019))，另一种是gasdermin D(GSDMD)或gasdermin E(GSDME)介导的细胞焦亡(参见Y.Wang等人，Chemotherapy drugs induce pyroptosis through caspase-3cleavage of agasdermin.Nature 547,99-103(2017)；和S.Ruhl等人ESCRT-dependent membrane repairnegatively regulates pyroptosis downstream of GSDMD activation.Science 362,956-960(2018))。其中，MLKL介导的细胞程序性坏死主要是通过TNF-a受体招募受体蛋白激酶1(RIP1)和3(RIP3)形成死亡复合物，随后激活MLKL以产生膜纳米孔，从而导致坏死性细胞死亡。与MLKL不同的是，GSDMD或GSDME由炎性半胱天冬酶(半胱天冬酶-1、-4、-5和-11)或半胱天冬酶-3激活，可以形成寡聚物并插入细胞膜中形成孔，因此介导细胞焦亡。
为了克服现有技术中CAR T治疗肿瘤时由于细胞因子释放综合征而引起的不良后果，需要新的策略在维持或提高CAR T细胞疗法功效的同时，控制细胞因子释放综合征，尤其是细胞因子风暴引起的安全性的问题。本发明人通过阐述两种新的细胞程序性死亡与CRS之间的关系，为CAR-T诱导的CRS的预测、治疗以及CAR-T细胞的临床应用提供了新的思路。
发明内容
我们的研究表明，在患有表达CD19+的恶性B细胞肿瘤中(例如Raji和NALM-6细胞)普遍表达GSDME。此外，在表达HER2+(例如SGC-7901和MCF-7)的肿瘤细胞也表达高水平的GSDME。我们进一步分析了人B细胞白血病患者中分离的原发性B-ALL白血病细胞中的GSDME水平与CD19-CAR T治疗后发生的CRS等级的相关性。根据我们的研究结果，提出以下实施方案。
一方面，本发明提供了特异性检测GSDME蛋白质或基因活性或水平的试剂在制备用于预测受试者发生细胞因子释放综合征的风险的试剂盒中的用途；优选地，所述试剂用于检测GSDME蛋白质或mRNA的表达水平。
在本发明提供的用途中，如果受试者GSDME蛋白质或基因活性或水平高于参照活性或水平，则预测所述受试者具有发生细胞因子释放综合征的风险，优选地，如果受试者GSDME蛋白质或基因活性或水平高于参照活性或水平的2倍，预测所述受试者具有发生严重细胞因子释放综合征的风险。优选地，根据本发明，严重细胞因子释放综合征是指根据临床分类的IV级(表现为威胁生命的症状并需要呼吸机支持)和V级(死亡)的细胞因子释放综合征。
在本发明一个特定的实施方案中，GSDME蛋白质或基因的参照活性或水平是指预先确定的在CAR T细胞治疗后不发生细胞因子释放综合征的患者中的GSDME蛋白质或基因的活性或水平。
在优选的实施方案中，参照活性或水平是预先确定的不发生细胞因子释放综合征的受试者中的GSDME蛋白质或基因活性或水平。
根据本发明所述的用途，其中所述受试者是患有癌症的受试者；优选地，所述癌症与CD19和/或HER2表达相关；优选地，所述受试者患有选自以下的癌症：B细胞急性淋巴细胞白血病(B-ALL)、T细胞急性淋巴细胞白血病(T-ALL)、急性淋巴细胞白血病(ALL)、慢性骨髓性白血病(CML)、慢性淋巴细胞性白血病(CLL)、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤(MCL)、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
在进一步的实施方案中，GSDME蛋白质或基因分离自受试者的癌症细胞，优选地，分离自受试者的表达CD19和/或HER2的癌细胞；进一步优选地，分离自选自以下的癌症的细胞：B细胞急性淋巴细胞白血病(B-ALL)、T细胞急性淋巴细胞白血病(T-ALL)、急性淋巴细胞白血病(ALL)、慢性骨髓性白血病(CML)、慢性淋巴细胞性白血病(CLL)、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤(MCL)、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
在另一个实施方案中，提供了在接受CAR T细胞疗法之前评估患有癌症的受试者发生细胞因子释放综合征的风险的方法，所述方法包括检测从受试者的癌细胞中分离的GSDME蛋白质或基因活性或水平，优选地，所述CAR T细胞为CD19和/或HER2 CAR T细胞。
另一方面，本发明提供了阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂在制备用于抑制和/或降低受试者发生细胞因子释放综合征的药物中的用途；优选地，所述受试者是癌症患者，更优选地，所述受试者患有与CD19和/或HER2表达相关的癌症；优选地，所述受试者患有选自以下的癌症：B细胞急性淋巴细胞白血病(B-ALL)、T细胞急性淋巴细胞白血病(T-ALL)、急性淋巴细胞白血病(ALL)、慢性骨髓性白血病(CML)、慢性淋巴细胞性白血病(CLL)、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤(MCL)、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
在进一步优选的实施方案中，阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂用于抑制和/或降低CAR T细胞治疗(例如，CD19和/或HER2 CAR T细胞治疗)引起的细胞因子释放综合征。
根据本发明，阻断和/或抑制GSDME蛋白质或基因活性或水平是指降低、减少、抑制或消除GSDME蛋白质或基因，例如敲除GSDME基因、降低mRNA的活性或表达水平。
已知激活的半胱天冬酶-3可以裂解GSDME产生其活性形式，该活性形式插入细胞膜中导致孔形成和随后的细胞焦亡(Y.Wang等人，Chemotherapy drugs inducepyroptosis through caspase-3 cleavage of a gasdermin.Nature 547,99-103(2017)；)。
因此，在一个优选的实施方案中，本发明所述的阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂选自抑制GSDME蛋白质活性的试剂(例如半胱天冬酶-1、-3、-4、-5、或11的抑制剂)、GSDME蛋白拮抗剂、抗-GSDME抗体、以及使GSDME基因(例如mRNA或DNA)表达缺失或降低的试剂(例如GSDME mRNA的反义序列例如miRNA，用于通过CRISPR-Cas9敲除GSDMEDNA的试剂)；在更优选的实施方案中，所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂是半胱天冬酶-3抑制剂，更优选地，所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂是贝纳卡桑(belnacasan)。
另一方面，本发明提供了一种治疗癌症的组合物，其包含CAR T(例如，CD19 CAR T细胞、或HER2 CAR T细胞)和一种或多种阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂，优选地，所述的阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂选自抑制GSDME蛋白质活性的试剂(例如半胱天冬酶-1、-3、-4、-5、或11的抑制剂)、GSDME蛋白拮抗剂、抗-GSDME抗体、以及使GSDME基因(例如mRNA或DNA)表达缺失或降低的试剂(例如GSDME mRNA的反义序列例如miRNA，用于通过CRISPR-Cas9敲除GSDME DNA的试剂)。在更优选的实施方案中，所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂是半胱天冬酶-3抑制剂，更优选地，所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂是贝纳卡桑(belnacasan)。优选地，所述癌症为与CD19和/或HER2表达相关的癌症；优选地，所述癌症选自：B细胞急性淋巴细胞白血病(B-ALL)、T细胞急性淋巴细胞白血病(T-ALL)、急性淋巴细胞白血病(ALL)、慢性骨髓性白血病(CML)、慢性淋巴细胞性白血病(CLL)、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤(MCL)、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
根据本发明提供的组合物，其中所述CAR T(例如，CD19 CAR T细胞、或HER2 CAR T细胞)和一种或多种阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂可以同时、分开或相继施用。
根据本发明提供的组合物可以有效降低CAR T细胞治疗引起的细胞因子综合征，而不改变CAR T细胞对肿瘤细胞的杀伤效果。
附图说明
图1A至图1G：HER2 CAR T或者CD19 CAR T细胞与Raji细胞或者NALM-6细胞按照不同效靶比共孵育6小时或者按照2:1的效靶比处理不同的时间点，然后检测细胞膜联蛋白V单阳以及和膜联蛋白V和PI双阳细胞比例。同时检测上清中LDH的释放量。
图2A至图2G：HER2 CAR T或者CD19 CAR T细胞与野生型Raji细胞或者NALM-6细胞或者GSDME敲除Raji细胞或者NALM-6细胞按照2:1的效靶比处理6个小时，然后检测细胞膜联蛋白V单阳以及和膜联蛋白V和PI双阳细胞比例。同时检测上清中LDH的释放量。
图3A至图3D：野生型Raji细胞或者NALM-6细胞或者GSDME敲除Raji细胞或者NALM-6细胞通过尾静脉接种于免疫缺陷鼠体内，21天后尾静脉注射10倍于肿瘤细胞的量的CAR T细胞，检测注射前和注射48小时后血清中炎症因子SAA、IL-6和IL-1β的含量以及小鼠的生存期。
图4A至图4D：野生型Raji细胞或者NALM-6细胞通过尾静脉接种于免疫缺陷鼠体内，21天后尾静脉注射10倍于肿瘤细胞的量的CAR T细胞，同时在CAR T回输前一周尾静脉注射巨噬细胞清除剂氯弗松-A(clophosome-A)或者半胱天冬酶-1抑制剂贝纳卡桑(belnacasan)，检测注射前和注射48小时后血清中炎症因子SAA、IL-6和IL-1β的含量以及小鼠的生存期。
图5A表示对人B-ALL患者中分离出的原代B白血病细胞(n＝11)的裂解物进行的抗GSDME的蛋白质印迹分析；
图5B表示人B-ALL患者中原代B白血病细胞的GSDME表达与CRS评分之间的相关性(n＝11)；
图5C表示人B-ALL患者中血清LDH水平；
图5D表示人B-ALL患者中血清LDH水平与CRS级别的相关性。
具体实施方式
该研究的主要目的是阐明CRS发生的潜在机制。针对复发或难治性B细胞急性淋巴母细胞白血病(R/R ALL)，离体产生了CD19 CAR T和HER2 CAR T细胞，使用CD19 TCR-ζ/4-1BB慢病毒载体转导的自体T细胞，体外表达含有CD3ζ结构域的CAR，以提供T细胞激活信号和4-1BB结构域，以提供共刺激信号。
我们还分析了人B细胞白血病患者中分离的原发性B-ALL白血病细胞中的GSDME水平与CD19-CAR T治疗后发生的CRS等级的相关性。
在体外研究中，使用CAR T或未修饰的T细胞与不同的肿瘤细胞共培养以确定肿瘤细胞的焦亡或凋亡。
另一方面，我们还使用了Cas9技术来敲除不同的基因，以阐明在小鼠模型中如何通过巨噬细胞依赖性途径激发焦亡引发的CRS。
实施例1.构建人CD19或HER2 CAR T细胞
HER2和CD19的嵌合抗原受体的构建如先前所述(R.A.Morgan,等人,Case reportof a serious adverse event following the administration of T cells transducedwith a chimeric antigen receptor recognizing ERBB2.Mol Ther 18,843-851(2010)；和M.C.Milone,等人，Chimeric receptors containing CD137 signal transductiondomains mediate enhanced survival of T cells and increased antileukemicefficacy in vivo.Mol Ther 17,1453-1464(2009))。简而言之，将来自mAb 4D5的HER2的单链Fv片段或来自克隆FMC63的CD19的单链Fv片段与具有CD3ζ和CD28细胞内信号传导结构域的CD8α链铰链和跨膜区连接，并将此盒插入慢病毒载体(Obioo Bioscience Company提供)。最后根据制造商的说明，用CD3/CD28激活珠(Invitrogen)刺激CD8+T细胞，从而启动转导，其中在含5％FBS的Vivo-15培养基(Lonza)中重组人IL-2的终浓度为100U/ml。在第2天收获细胞用于慢病毒转导，并重悬于相同培养基中。将含有慢病毒的上清液以MOI 1:10的比例添加到培养基中，并根据生产商的说明，用RetroNectin(CH-296；Takara Bio，Ohtsu，日本，用10mg/ml CH-296进行包被)包被平板。然后，将细胞在32℃下以1000g离心2小时，并在37℃下孵育6小时。2天后用流式细胞仪定量感染率。在这项研究中，根据效应细胞与靶细胞的比例(2：1)或体外实验中的指示，使用CD19-CAR T细胞的数量。用慢病毒-CAR转染T细胞，并在体内实验培养10天。在慢病毒转导后第3天和第5天，以及在培养结束时，通过流式细胞术评估转染效率。转染效率约为40％。对于体外实验，我们培养CAR T细胞5-7天。这些T细胞在扩增期间对数生长。对于临床试验，我们使用CD3ζ-4-1BB-CART。否则，使用CD3ζ-CD28-CART细胞。
实施例2.构建具有人CD19的小鼠CAR T(mCAR hCD19)
mCAR-hCD19的序列包含人CD19的抗原受体或HER2单链可变片段、鼠CD3ζ、CD28和/或4-1-BB以及N末端的myc标签，如前所述(参见，例如J.Chen等人,NR4A transcriptionfactors limit CAR T cell function in solid tumours.Nature 567,530-534(2019))，其由SyngenTech合成。然后将该嵌合抗原构建体克隆到MSCV-GFP(Clontech)鼠逆转录病毒载体(MSCV-myc-CAR-2A)中。然后，用mCAR-hCD19质粒和pCL-Eco逆转录病毒包装质粒转染Platinum-E逆转录病毒包装细胞系Ecotropic(PlatE)细胞(Cell Biolabs，RV-101)，以获得含有mCAR-hCD19的逆转录病毒。OT-1CD8+T细胞被抗CD3/CD28磁珠(Gibco，11453D)、IL-2(Perprotech，212-12)和55μMβ-巯基乙醇(Gibco，21985-023)激活，持续24小时。然后在RetroNectin(Takara Bio)存在下，用以上病毒感染OT-1CD8+T细胞8小时。24小时后，使用BD Biosciences FACSAria II通过流式细胞仪分选GFP阳性细胞，获得表达高水平的hCD19或hHER2的细胞。
实施例3.CAR T细胞体外诱导肿瘤细胞焦亡
将如上实施例1中制备的HER2 CAR T或者CD19 CAR T细胞与Raji细胞或者NALM-6细胞按照不同效靶比共孵育6小时或者按照2:1的效靶比处理不同的时间点，通过流式细胞术检测细胞膜联蛋白V单阳以及和膜联蛋白V和PI双阳细胞比例(参见图1A至图1C)。并使用ELISA试剂盒(eBiosence,CA,USA)根据制造商的说明书检测上清液中的LDH的释放量(参见图1D至图1G)。通过学生t检验或单向方差分析，**p<0.01，***p<0.001。数据代表三个独立实验的平均值±SD。
由上述结果表明，CAR T在杀伤肿瘤细胞时引起的是焦亡，而非凋亡，由膜联蛋白V和PI双阳细胞比例升高以及LDH释放量增加可知。
实施例4.通过CRISPR-Cas9产生稳定的GSDME基因敲除细胞系
为了产生稳定的GSDME敲除的Raji细胞或者NALM-6细胞系，采用本领域熟知的CRISPR-Cas9方法敲除Raji细胞或者NALM-6细胞系中的GSDME基因。简言之，将GSDME基因对应别的SGRNA克隆到pL-CRISPR.EFS.GFP载体质粒中，并包装质粒psPAX2和pMD2.G共同转染HEK 293T细胞。48小时后，收获慢病毒并浓缩，与终浓度为8μg/ml的聚乙烯共同感染Raji和NALM-6细胞系。两天后，通过蛋白质印迹杂交确认基因敲除的细胞。
实施例5.CAR T细胞体外诱导肿瘤细胞焦亡是通过GSDME介导的HER2 CAR T或者CD19 CAR T细胞与野生型Raji细胞或者NALM-6细胞细胞或者GSDME敲除的Raji细胞或者NALM-6细胞按照2:1的效靶比处理6个小时，然后通过流式细胞术检测细胞膜联蛋白V单阳以及和膜联蛋白V和PI双阳细胞比例(参见图2A至图2E)。同时使用ELISA试剂盒(eBiosence,CA,USA)根据制造商的说明书检测上清中LDH的释放量(参见图2F至图2G)。通过学生t检验或单向方差分析，**p<0.01，***p<0.001。数据代表三个独立实验的平均值±SD。
由图2A和2G可知，GSDME的敲除后是CAR T杀伤肿瘤细胞时引起的焦亡转换为凋亡，由膜联蛋白V和PI双阳细胞比例升高以及LDH释放量增加可知；但不改变杀伤效果，因为膜联蛋白V单阳细胞比例没有降低。
实施例6.CAR T细胞治疗在小鼠体内诱发肿瘤细胞焦亡和CRS
将野生型Raji细胞或者NALM-6细胞或者GSDME敲除的Raji细胞或者NALM-6细胞通过尾静脉接种于免疫缺陷鼠SCID-beige体内，21天后检测肿瘤负荷，符合要求的小鼠尾静脉注射10倍于肿瘤细胞的量的CAR T细胞。在CAR T细胞注射前4小时和注射后48小时使用ELISA试剂盒(eBiosence,CA,USA)根据制造商的说明书检测小鼠血清中炎症因子SAA、IL-6和IL-1β的含量(参见图3A至图3C)，并记录小鼠的生存期(参见图3D)。通过学生t检验或单向方差分析，**p<0.01，***p<0.001。数据代表三个独立实验的平均值±SD。
其结果表明，GSDME的敲除能缓解CAR T引起的炎症因子释放(见，图3A至图3C)，减轻由于CRS引起的死亡，延长存活(图3D)。
实施例7.巨噬细胞清除剂或半胱天冬酶-1抑制剂减轻CAR T细胞治疗在小鼠体内诱发的CRS
野生型Raji细胞或者NALM-6细胞通过尾静脉接种于免疫缺陷小鼠SCID-beige体内，21天后检测肿瘤负荷，符合要求的小鼠尾静脉注射10倍于肿瘤细胞的量的CAR T细胞，在CAR T细胞注射前一周尾静脉注射巨噬细胞清除剂氯弗松-A(clophosome-A)(200μl/只小鼠)或者半胱天冬酶-1抑制剂贝纳卡桑(belnacasan)(100mg/kg)，每隔一天一次，共3次。在CAR T细胞检测注射前和注射48小时后使用ELISA试剂盒(eBiosence,CA,USA)根据制造商的说明书检测血清中炎症因子SAA、IL-6和IL-1β的含量(图4A至图4C)，并记录小鼠的生存期(图4D)。通过学生t检验或单向方差分析，**p<0.01，***p<0.001。数据代表三个独立实验的平均值±SD。
结果表明，巨噬细胞清除剂氯弗松-A或者半胱天冬酶-1抑制剂贝纳卡桑能缓解CAR T引起的炎症因子释放(参见图4A至图4C)并减轻由于CRS引起的死亡(参见图4D)。这与我们的研究结果肿瘤细胞焦亡触发巨噬细胞释放促炎性细胞因子一致。
实施例8.人B细胞白血病患者中原发性B-ALL白血病细胞的GSDME表达水平与CD19-CAR T诱导的CRS等级的相关性。
患者来自郑州大学第一附属医院，为了符合参加研究的资格，筛查年龄至少4岁且不超过70岁、并且被诊断为CD19+复发或难治性B细胞白血病的患者，患者被诊断为不符合自体或异源性SCT，且根据目前可用的疗法预后有限(数月至<2年生存期)；ECOG结果为0、1或2；具有稳定的生命体征。其他合格标准是心脏、肝脏和肾脏功能健全。所有患者均提供书面知情同意书以参加研究。从这些患者获得外周血或骨髓。实验获得郑州大学第一附属医院临床试验伦理委员会的伦理许可。
细胞因子释放综合征(CRS)等级
CRS的等级系统是根据临床分类进行的(参见，S.S.Neelapu等人，Chimericantigen receptor T-cell therapy—assessment and management of toxicities.NatRev Clin Oncol 15,47(2017))。简而言之，I级为并不威胁生命的症状，例如发烧、头痛、肌痛、不适、恶心或疲劳等；II级包括需要静脉输液或小剂量血管加压药、或对其有反应的症状，II级的器官毒性或吸氧分数低于40％；III级包括需要积极干预(大剂量或多种升压药)或对其有反应的症状，III级的器官毒性或吸氧分数等于或超过40％；IV级表现为威胁生命的症状，需要呼吸机支持；V级为死亡。器官毒性根据CTCAE v4.03进行分级(参见CommonTerminology Criteria for Adverse Events(CTCAE)Version 5.0.(NIH Publication,2017).U.S.DEPARTMENT OF HEALTH AND HUMAN 12SERVICES)。
从患者分离B细胞白血病细胞，在M2裂解液中裂解，超声波处理。采用BCA试剂盒(Applygen Technologies Inc.,China)对蛋白质定量。然后，在SDS-PAGE凝胶上分离蛋白质，并转移到硝化纤维素膜上。在5％BSA中封闭膜，并与抗GSDME(Abcam,UK)抗体过夜孵育，然后与偶联辣根过氧化物酶的二抗孵育，并检测。
我们分析了CD19-CAR T治疗前从11例患者中分离出的原发性B-ALL白血病细胞中的GSDME水平。尽管来自患者的B白血病细胞普遍表达GSDME(图5A)，但在这些患者中，更高水平的GSDME与更严重的CRS病例相关(图5B)。
另外，我们使用ELISA试剂盒(eBiosence,CA,USA)根据制造商的说明书检测了CART治疗后患者的血清LDH水平。
发现CRS等级较高的患者(n＝7)与低CRS(n＝4)相比，血LDH水平)为更高(图5C)，而LDH的水平与CRS的严重程度呈正相关(图5D)。这些在一起表明，CAR T细胞疗法诱导的B白血病细胞焦亡引发了患者的CRS。
Claims (10)
1.特异性检测GSDME蛋白质或基因活性或水平的试剂在制备用于预测受试者发生细胞因子释放综合征的风险的试剂盒中的用途；优选地，所述试剂用于检测GSDME蛋白质或mRNA的表达水平。
2.根据权利要求1所述的用途，其中如果受试者GSDME蛋白质或基因活性或水平高于参照活性或水平，则预测所述受试者具有发生细胞因子释放综合征的风险，优选地，如果受试者GSDME蛋白质或基因活性或水平高于参照活性或水平的2倍，预测所述受试者具有发生严重细胞因子释放综合征的风险。
3.根据权利要求1或2所述的用途，其中所述受试者是患有癌症的受试者；优选地，所述癌症与CD19和/或HER2表达相关；优选地，所述受试者患有选自以下的癌症：B细胞急性淋巴细胞白血病、T细胞急性淋巴细胞白血病、急性淋巴细胞白血病、慢性骨髓性白血病、慢性淋巴细胞性白血病、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
4.根据权利要求1或2所述的用途，其中在接受CAR T细胞疗法之前评估受试者发生细胞因子释放综合征的风险；优选地，所述CAR T细胞为CD19和/或HER2 CAR T细胞。
5.阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂在制备用于抑制和/或降低受试者发生细胞因子释放综合征的药物中的用途，优选地，所述受试者患有癌症，更优选地，所述受试者患有与CD19和/或HER2表达相关的癌症；优选地，所述受试者患有选自以下的癌症：B细胞急性淋巴细胞白血病、T细胞急性淋巴细胞白血病、急性淋巴细胞白血病、慢性骨髓性白血病、慢性淋巴细胞性白血病、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
6.根据权利要求5所述的用途，其中所述的阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂选自抑制GSDME蛋白质活性的试剂、降低GSDME蛋白质表达水平的试剂、降低GSDME mRNA表达水平的试剂和敲除GSDME DNA的试剂；优选地，所述抑制GSDME蛋白质活性的试剂选自半胱天冬酶-1、-3、-4、-5、或11的抑制剂；所述降低GSDME蛋白质表达水平的试剂优选选自GSDME蛋白拮抗剂和抗-GSDME抗体；所述降低GSDME mRNA表达水平的试剂优选选自GSDME mRNA的反义序列；所述敲除GSDME DNA的试剂优选选自用于通过CRISPR-Cas9敲除GSDME DNA的试剂。
7.根据权利要求5或6所述的用途，其中所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂用于抑制和/或降低CAR T细胞治疗引起的细胞因子释放综合征。
8.一种用于治疗癌症的组合物，其中所述组合物包含CAR T和一种或多种阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂，优选地，所述阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂选自抑制GSDME蛋白质活性的试剂、降低GSDME蛋白质表达水平的试剂、降低GSDME mRNA表达水平的试剂和敲除GSDME DNA的试剂；优选地，所述抑制GSDME蛋白质活性的试剂选自半胱天冬酶-1、-3、-4、-5、或11的抑制剂；所述降低GSDME蛋白质表达水平的试剂优选选自GSDME蛋白拮抗剂和抗-GSDME抗体；所述降低GSDME mRNA表达水平的试剂优选选自GSDME mRNA的反义序列；所述敲除GSDME DNA的试剂优选选自用于通过CRISPR-Cas9敲除GSDME DNA的试剂。
9.根据权利要求8所述的组合物在制备用于治疗受试者癌症的药物中的用途，优选地，所述癌症为与CD19和/或HER2表达相关的癌症；优选地，所述癌症选自：B细胞急性淋巴细胞白血病、T细胞急性淋巴细胞白血病、急性淋巴细胞白血病、慢性骨髓性白血病、慢性淋巴细胞性白血病、B细胞早幼粒细胞白血病、胚细胞性浆细胞样树突状细胞瘤、伯基特淋巴瘤、弥漫性大B细胞淋巴瘤、滤泡淋巴瘤、毛细胞白血病、小细胞或大细胞滤泡淋巴瘤、恶性淋巴细胞增生症、MALT淋巴瘤、外套细胞淋巴瘤、边缘区淋巴瘤、多发性骨髓瘤、骨髓发育不良和骨髓增生异常综合征、非霍奇金淋巴瘤、霍奇金淋巴瘤、浆胚细胞淋巴瘤、浆细胞样树突状细胞肿瘤和瓦尔登斯特隆巨球蛋白血症。
10.根据权利要求9所述的用途，其中所述组合物中的CAR T和一种或多种阻断和/或抑制GSDME蛋白质或基因活性或水平的试剂同时、分开或相继施用。
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010035350.7A CN113189340A (zh) | 2020-01-14 | 2020-01-14 | 细胞焦亡通路在细胞治疗中的用途 |
| PCT/CN2020/137498 WO2021143455A1 (zh) | 2020-01-14 | 2020-12-18 | 细胞焦亡通路在细胞治疗中的用途 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010035350.7A CN113189340A (zh) | 2020-01-14 | 2020-01-14 | 细胞焦亡通路在细胞治疗中的用途 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113189340A true CN113189340A (zh) | 2021-07-30 |
Family
ID=76863557
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010035350.7A Pending CN113189340A (zh) | 2020-01-14 | 2020-01-14 | 细胞焦亡通路在细胞治疗中的用途 |
Country Status (2)
| Country | Link |
|---|---|
| CN (1) | CN113189340A (zh) |
| WO (1) | WO2021143455A1 (zh) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115819548B (zh) * | 2021-11-16 | 2023-09-01 | 北京美德泰康生物科技有限公司 | 一种检测炎症相关疾病的标志物和方法 |
| LU501764B1 (en) | 2022-03-31 | 2023-10-02 | Leibniz Inst Fuer Naturstoff Forschung Und Infektionsbiologie E V Hans Knoell Inst Hki | Gasdermin e expression in human t cells as a marker for proinflammatory t cell functions |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20180296572A1 (en) * | 2017-04-01 | 2018-10-18 | Avm Biotechnology, Llc | Replacement of cytotoxic preconditioning before cellular immunotherapy |
| CN109069539A (zh) * | 2016-02-18 | 2018-12-21 | 恩立夫克治疗有限责任公司 | 用于癌症治疗的联合免疫疗法和细胞因子控制疗法 |
| CN110582287A (zh) * | 2017-02-27 | 2019-12-17 | 朱诺治疗学股份有限公司 | 与在细胞疗法中给药有关的组合物、制品和方法 |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109564225B (zh) * | 2016-08-09 | 2022-05-13 | B.R.A.H.M.S 有限公司 | 作为指示不良事件的标志物的组蛋白和/或proADM |
| US20210046159A1 (en) * | 2018-03-09 | 2021-02-18 | Ospedale San Raffaele S.R.L. | Il-1 antagonist and toxicity induced by cell therapy |
| GB201804514D0 (en) * | 2018-03-21 | 2018-05-02 | Ucl Business Plc | Treatment of pyroptosis |
| US20210267996A1 (en) * | 2018-06-27 | 2021-09-02 | Children`S Medical Center Corporation | Compounds for inhibition of inflammation |
| CN108721629A (zh) * | 2018-07-17 | 2018-11-02 | 厦门大学 | 一种抗肿瘤药物组合物及其应用包含铁离子的试剂 |
| CN110664804B (zh) * | 2019-10-12 | 2022-08-26 | 南京大学 | 化合物ag-041r在制备抵抗免疫细胞焦亡药物中的应用 |
-
2020
- 2020-01-14 CN CN202010035350.7A patent/CN113189340A/zh active Pending
- 2020-12-18 WO PCT/CN2020/137498 patent/WO2021143455A1/zh active Application Filing
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109069539A (zh) * | 2016-02-18 | 2018-12-21 | 恩立夫克治疗有限责任公司 | 用于癌症治疗的联合免疫疗法和细胞因子控制疗法 |
| CN110582287A (zh) * | 2017-02-27 | 2019-12-17 | 朱诺治疗学股份有限公司 | 与在细胞疗法中给药有关的组合物、制品和方法 |
| US20180296572A1 (en) * | 2017-04-01 | 2018-10-18 | Avm Biotechnology, Llc | Replacement of cytotoxic preconditioning before cellular immunotherapy |
Non-Patent Citations (1)
| Title |
|---|
| YUYING LIU等: "Gasdermin E-mediated target cell pyroptosis by CAR T cells triggers cytokine release syndrome", 《SCI IMMUNOL.》 * |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2021143455A1 (zh) | 2021-07-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Pellegatta et al. | Constitutive and TNFα-inducible expression of chondroitin sulfate proteoglycan 4 in glioblastoma and neurospheres: Implications for CAR-T cell therapy | |
| Zhao et al. | Engineered IL-7 receptor enhances the therapeutic effect of AXL-CAR-T cells on triple-negative breast cancer | |
| Faust et al. | Correlation of renal tubular epithelial cell–derived interleukin‐18 up‐regulation with disease activity in MRL‐Faslpr mice with autoimmune lupus nephritis | |
| US20190077856A1 (en) | Method of treating diseases using kinase modulators | |
| Tsujimura et al. | Transcriptional regulation of multidrug resistance‐1 gene by interleukin‐2 in lymphocytes | |
| Dey et al. | Peptide vaccination directed against IDO1-expressing immune cells elicits CD8+ and CD4+ T-cell-mediated antitumor immunity and enhanced anti-PD1 responses | |
| WO2021143455A1 (zh) | 细胞焦亡通路在细胞治疗中的用途 | |
| Uota et al. | An IκB kinase 2 inhibitor IMD‐0354 suppresses the survival of adult T‐cell leukemia cells | |
| Yang et al. | PD-1, PD-L1 and PD-L2 expression in mouse prostate cancer | |
| Hong et al. | Gal-3BP negatively regulates NF-κB signaling by inhibiting the activation of TAK1 | |
| KR102502736B1 (ko) | 항암 및 면역증강용 신규 타겟 | |
| Wei et al. | Low tristetraprolin expression promotes cell proliferation and predicts poor patients outcome in pancreatic cancer | |
| Peled et al. | EF hand domain family member D2 is required for T cell cytotoxicity | |
| Lou et al. | N-cadherin protects oral cancer cells from NK cell killing in the circulation by inducing NK cell functional exhaustion via the KLRG1 receptor | |
| Yang et al. | CD44 v5 domain inhibition represses the polarization of Th2 cells by interfering with the IL‐4/IL‐4R signaling pathway | |
| Ba et al. | Antitumor effect of CAR-T cells targeting transmembrane tumor necrosis factor alpha combined with PD-1 mAb on breast cancers | |
| Vasilevsky et al. | OX40 engagement stabilizes Mxd4 and Mnt protein levels in antigen‐stimulated T cells leading to an increase in cell survival | |
| Wang et al. | TIGIT immune checkpoint blockade enhances immunity of human peripheral blood NK cells against castration-resistant prostate cancer | |
| Serrano‐Pertierra et al. | Wiskott‐Aldrich syndrome protein (WASp) and N‐WASp are involved in the regulation of NK‐cell migration upon NKG2D activation | |
| Hamilton et al. | Loss of the cyclin-dependent kinase inhibitor 1 in the context of brachyury-mediated phenotypic plasticity drives tumor resistance to immune attack | |
| US20200323952A1 (en) | Compositions and methods of treating cancer | |
| Huang et al. | Inhibition of Fas associated phosphatase 1 (Fap1) facilitates apoptosis of colon cancer stem cells and enhances the effects of oxaliplatin | |
| US20230357388A1 (en) | Immunotherapy | |
| Hsu et al. | Role for a zinc finger protein (Zfp111) in transformation of 208F rat fibroblasts by jaagsiekte sheep retrovirus envelope protein | |
| Huang et al. | Checkpoint CD24 function on tumor and immunotherapy |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210730 |
|
| RJ01 | Rejection of invention patent application after publication |