WO2020018727A1 - T cell-antigen coupler with various construct optimizations - Google Patents
T cell-antigen coupler with various construct optimizations Download PDFInfo
- Publication number
- WO2020018727A1 WO2020018727A1 PCT/US2019/042297 US2019042297W WO2020018727A1 WO 2020018727 A1 WO2020018727 A1 WO 2020018727A1 US 2019042297 W US2019042297 W US 2019042297W WO 2020018727 A1 WO2020018727 A1 WO 2020018727A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- seq
- acid sequence
- nucleic acid
- tac
- domain
- Prior art date
Links
- 239000000427 antigen Substances 0.000 title claims description 158
- 238000005457 optimization Methods 0.000 title 1
- 239000003446 ligand Substances 0.000 claims abstract description 340
- 108091008874 T cell receptors Proteins 0.000 claims abstract description 271
- 102000016266 T-Cell Antigen Receptors Human genes 0.000 claims abstract description 269
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract description 103
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 94
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 94
- 229940127174 UCHT1 Drugs 0.000 claims abstract description 90
- 102000004196 processed proteins & peptides Human genes 0.000 claims abstract description 84
- 229920001184 polypeptide Polymers 0.000 claims abstract description 83
- 230000011664 signaling Effects 0.000 claims abstract description 53
- 210000001744 T-lymphocyte Anatomy 0.000 claims description 268
- 206010028980 Neoplasm Diseases 0.000 claims description 226
- 150000007523 nucleic acids Chemical group 0.000 claims description 198
- 230000001086 cytosolic effect Effects 0.000 claims description 191
- 108091033319 polynucleotide Proteins 0.000 claims description 178
- 102000040430 polynucleotide Human genes 0.000 claims description 178
- 239000002157 polynucleotide Substances 0.000 claims description 178
- 210000004027 cell Anatomy 0.000 claims description 156
- 108091028043 Nucleic acid sequence Proteins 0.000 claims description 143
- 108091007433 antigens Proteins 0.000 claims description 139
- 102000036639 antigens Human genes 0.000 claims description 139
- 101001012157 Homo sapiens Receptor tyrosine-protein kinase erbB-2 Proteins 0.000 claims description 93
- 102100030086 Receptor tyrosine-protein kinase erbB-2 Human genes 0.000 claims description 93
- 101000716102 Homo sapiens T-cell surface glycoprotein CD4 Proteins 0.000 claims description 89
- 102100036011 T-cell surface glycoprotein CD4 Human genes 0.000 claims description 89
- 101000946843 Homo sapiens T-cell surface glycoprotein CD8 alpha chain Proteins 0.000 claims description 82
- 102100034922 T-cell surface glycoprotein CD8 alpha chain Human genes 0.000 claims description 82
- 108700022150 Designed Ankyrin Repeat Proteins Proteins 0.000 claims description 78
- 201000011510 cancer Diseases 0.000 claims description 78
- 239000013598 vector Substances 0.000 claims description 56
- 108010008014 B-Cell Maturation Antigen Proteins 0.000 claims description 55
- 102000006942 B-Cell Maturation Antigen Human genes 0.000 claims description 55
- 230000027455 binding Effects 0.000 claims description 53
- 238000000034 method Methods 0.000 claims description 44
- 239000008194 pharmaceutical composition Substances 0.000 claims description 40
- 230000004913 activation Effects 0.000 claims description 32
- 230000035772 mutation Effects 0.000 claims description 21
- 239000012634 fragment Substances 0.000 claims description 18
- 208000024893 Acute lymphoblastic leukemia Diseases 0.000 claims description 12
- 206010035226 Plasma cell myeloma Diseases 0.000 claims description 12
- 108020001580 protein domains Proteins 0.000 claims description 12
- 208000005718 Stomach Neoplasms Diseases 0.000 claims description 11
- 206010017758 gastric cancer Diseases 0.000 claims description 11
- 201000011549 stomach cancer Diseases 0.000 claims description 11
- 210000004962 mammalian cell Anatomy 0.000 claims description 10
- 208000003950 B-cell lymphoma Diseases 0.000 claims description 9
- 102100024222 B-lymphocyte antigen CD19 Human genes 0.000 claims description 9
- 101000980825 Homo sapiens B-lymphocyte antigen CD19 Proteins 0.000 claims description 9
- 208000034578 Multiple myelomas Diseases 0.000 claims description 9
- 239000007788 liquid Substances 0.000 claims description 9
- 206010006187 Breast cancer Diseases 0.000 claims description 8
- 208000026310 Breast neoplasm Diseases 0.000 claims description 8
- 206010009944 Colon cancer Diseases 0.000 claims description 8
- 230000036210 malignancy Effects 0.000 claims description 8
- 208000010839 B-cell chronic lymphocytic leukemia Diseases 0.000 claims description 7
- 241000124008 Mammalia Species 0.000 claims description 7
- 206010033128 Ovarian cancer Diseases 0.000 claims description 7
- 206010061535 Ovarian neoplasm Diseases 0.000 claims description 7
- 108010049777 Ankyrins Proteins 0.000 claims description 6
- 102000008102 Ankyrins Human genes 0.000 claims description 6
- 208000015914 Non-Hodgkin lymphomas Diseases 0.000 claims description 6
- 210000003719 b-lymphocyte Anatomy 0.000 claims description 6
- 208000032839 leukemia Diseases 0.000 claims description 6
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 6
- 230000004936 stimulating effect Effects 0.000 claims description 6
- 239000002136 L01XE07 - Lapatinib Substances 0.000 claims description 5
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims description 5
- 229940018964 belantamab mafodotin Drugs 0.000 claims description 5
- 229950008579 ertumaxomab Drugs 0.000 claims description 5
- 229940121448 gancotamab Drugs 0.000 claims description 5
- 229960004891 lapatinib Drugs 0.000 claims description 5
- BCFGMOOMADDAQU-UHFFFAOYSA-N lapatinib Chemical compound O1C(CNCCS(=O)(=O)C)=CC=C1C1=CC=C(N=CN=C2NC=3C=C(Cl)C(OCC=4C=C(F)C=CC=4)=CC=3)C2=C1 BCFGMOOMADDAQU-UHFFFAOYSA-N 0.000 claims description 5
- 201000005202 lung cancer Diseases 0.000 claims description 5
- 208000020816 lung neoplasm Diseases 0.000 claims description 5
- 229950003135 margetuximab Drugs 0.000 claims description 5
- 229960005558 mertansine Drugs 0.000 claims description 5
- 229950008835 neratinib Drugs 0.000 claims description 5
- ZNHPZUKZSNBOSQ-BQYQJAHWSA-N neratinib Chemical compound C=12C=C(NC\C=C\CN(C)C)C(OCC)=CC2=NC=C(C#N)C=1NC(C=C1Cl)=CC=C1OCC1=CC=CC=N1 ZNHPZUKZSNBOSQ-BQYQJAHWSA-N 0.000 claims description 5
- 229960002087 pertuzumab Drugs 0.000 claims description 5
- 229940060249 timigutuzumab Drugs 0.000 claims description 5
- 229960000575 trastuzumab Drugs 0.000 claims description 5
- 208000001333 Colorectal Neoplasms Diseases 0.000 claims description 4
- 206010014733 Endometrial cancer Diseases 0.000 claims description 4
- 206010014759 Endometrial neoplasm Diseases 0.000 claims description 4
- 206010025323 Lymphomas Diseases 0.000 claims description 4
- 208000029742 colonic neoplasm Diseases 0.000 claims description 4
- 208000005017 glioblastoma Diseases 0.000 claims description 4
- 206010044412 transitional cell carcinoma Diseases 0.000 claims description 4
- 230000003442 weekly effect Effects 0.000 claims description 4
- 206010005003 Bladder cancer Diseases 0.000 claims description 3
- 206010061902 Pancreatic neoplasm Diseases 0.000 claims description 3
- 208000007097 Urinary Bladder Neoplasms Diseases 0.000 claims description 3
- 208000015486 malignant pancreatic neoplasm Diseases 0.000 claims description 3
- 201000002528 pancreatic cancer Diseases 0.000 claims description 3
- 208000008443 pancreatic carcinoma Diseases 0.000 claims description 3
- 239000007787 solid Substances 0.000 claims description 3
- 201000005112 urinary bladder cancer Diseases 0.000 claims description 3
- 125000003275 alpha amino acid group Chemical group 0.000 claims 19
- BGFTWECWAICPDG-UHFFFAOYSA-N 2-[bis(4-chlorophenyl)methyl]-4-n-[3-[bis(4-chlorophenyl)methyl]-4-(dimethylamino)phenyl]-1-n,1-n-dimethylbenzene-1,4-diamine Chemical compound C1=C(C(C=2C=CC(Cl)=CC=2)C=2C=CC(Cl)=CC=2)C(N(C)C)=CC=C1NC(C=1)=CC=C(N(C)C)C=1C(C=1C=CC(Cl)=CC=1)C1=CC=C(Cl)C=C1 BGFTWECWAICPDG-UHFFFAOYSA-N 0.000 claims 6
- 238000010361 transduction Methods 0.000 abstract description 7
- 230000026683 transduction Effects 0.000 abstract description 7
- 239000012636 effector Substances 0.000 abstract description 4
- 208000004605 Persistent Truncus Arteriosus Diseases 0.000 description 218
- 208000037258 Truncus arteriosus Diseases 0.000 description 218
- 150000001413 amino acids Chemical group 0.000 description 204
- 239000002773 nucleotide Substances 0.000 description 150
- 125000003729 nucleotide group Chemical group 0.000 description 150
- 241000699670 Mus sp. Species 0.000 description 81
- 235000018102 proteins Nutrition 0.000 description 77
- 108010019670 Chimeric Antigen Receptors Proteins 0.000 description 52
- 102000039446 nucleic acids Human genes 0.000 description 45
- 108020004707 nucleic acids Proteins 0.000 description 45
- 102000005962 receptors Human genes 0.000 description 36
- 108020003175 receptors Proteins 0.000 description 36
- 238000011282 treatment Methods 0.000 description 35
- 235000001014 amino acid Nutrition 0.000 description 32
- 229940024606 amino acid Drugs 0.000 description 32
- 238000001727 in vivo Methods 0.000 description 31
- 210000004881 tumor cell Anatomy 0.000 description 28
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 27
- 201000010099 disease Diseases 0.000 description 26
- 238000001994 activation Methods 0.000 description 25
- 102000004127 Cytokines Human genes 0.000 description 21
- 108090000695 Cytokines Proteins 0.000 description 21
- 239000000203 mixture Substances 0.000 description 21
- 230000016396 cytokine production Effects 0.000 description 20
- 230000006044 T cell activation Effects 0.000 description 19
- 230000003013 cytotoxicity Effects 0.000 description 19
- 231100000135 cytotoxicity Toxicity 0.000 description 19
- 230000000694 effects Effects 0.000 description 17
- 108700018351 Major Histocompatibility Complex Proteins 0.000 description 16
- 238000000338 in vitro Methods 0.000 description 16
- 230000020382 suppression by virus of host antigen processing and presentation of peptide antigen via MHC class I Effects 0.000 description 16
- 210000004369 blood Anatomy 0.000 description 15
- 239000008280 blood Substances 0.000 description 15
- 238000013456 study Methods 0.000 description 15
- 241001529936 Murinae Species 0.000 description 14
- 230000007774 longterm Effects 0.000 description 13
- 238000004519 manufacturing process Methods 0.000 description 12
- 230000001965 increasing effect Effects 0.000 description 11
- 239000007924 injection Substances 0.000 description 11
- 238000002347 injection Methods 0.000 description 11
- 239000004475 Arginine Substances 0.000 description 10
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 10
- 230000037396 body weight Effects 0.000 description 10
- 239000003112 inhibitor Substances 0.000 description 10
- 238000001990 intravenous administration Methods 0.000 description 10
- 239000000126 substance Substances 0.000 description 10
- 238000006467 substitution reaction Methods 0.000 description 10
- 230000006870 function Effects 0.000 description 9
- 238000001802 infusion Methods 0.000 description 9
- 150000002632 lipids Chemical class 0.000 description 9
- 230000004083 survival effect Effects 0.000 description 9
- 101000914514 Homo sapiens T-cell-specific surface glycoprotein CD28 Proteins 0.000 description 8
- 241000713666 Lentivirus Species 0.000 description 8
- 241000699666 Mus <mouse, genus> Species 0.000 description 8
- 102100027213 T-cell-specific surface glycoprotein CD28 Human genes 0.000 description 8
- 238000009472 formulation Methods 0.000 description 8
- 230000002688 persistence Effects 0.000 description 8
- 230000002829 reductive effect Effects 0.000 description 8
- 208000024891 symptom Diseases 0.000 description 8
- 239000003981 vehicle Substances 0.000 description 8
- 241001465754 Metazoa Species 0.000 description 7
- 241000700605 Viruses Species 0.000 description 7
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 7
- 235000018417 cysteine Nutrition 0.000 description 7
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 6
- 102100033725 Tumor necrosis factor receptor superfamily member 16 Human genes 0.000 description 6
- 230000000259 anti-tumor effect Effects 0.000 description 6
- 238000009826 distribution Methods 0.000 description 6
- 239000003937 drug carrier Substances 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 230000012010 growth Effects 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 230000004044 response Effects 0.000 description 6
- 239000000243 solution Substances 0.000 description 6
- 208000014697 Acute lymphocytic leukaemia Diseases 0.000 description 5
- 208000011691 Burkitt lymphomas Diseases 0.000 description 5
- 108020004414 DNA Proteins 0.000 description 5
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 5
- 108010002350 Interleukin-2 Proteins 0.000 description 5
- 102000017578 LAG3 Human genes 0.000 description 5
- 208000025205 Mantle-Cell Lymphoma Diseases 0.000 description 5
- 108010032605 Nerve Growth Factor Receptors Proteins 0.000 description 5
- 208000006664 Precursor Cell Lymphoblastic Leukemia-Lymphoma Diseases 0.000 description 5
- 108010076504 Protein Sorting Signals Proteins 0.000 description 5
- 239000011324 bead Substances 0.000 description 5
- 230000010261 cell growth Effects 0.000 description 5
- 108700010039 chimeric receptor Proteins 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 230000018109 developmental process Effects 0.000 description 5
- 230000002147 killing effect Effects 0.000 description 5
- 210000004698 lymphocyte Anatomy 0.000 description 5
- 201000001441 melanoma Diseases 0.000 description 5
- 230000004048 modification Effects 0.000 description 5
- 238000012986 modification Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 230000007115 recruitment Effects 0.000 description 5
- 230000000638 stimulation Effects 0.000 description 5
- 230000008685 targeting Effects 0.000 description 5
- 230000001225 therapeutic effect Effects 0.000 description 5
- 230000001988 toxicity Effects 0.000 description 5
- 231100000419 toxicity Toxicity 0.000 description 5
- 230000004614 tumor growth Effects 0.000 description 5
- 239000013603 viral vector Substances 0.000 description 5
- 102100023635 Alpha-fetoprotein Human genes 0.000 description 4
- 108010022366 Carcinoembryonic Antigen Proteins 0.000 description 4
- 102100025475 Carcinoembryonic antigen-related cell adhesion molecule 5 Human genes 0.000 description 4
- 101000851370 Homo sapiens Tumor necrosis factor receptor superfamily member 9 Proteins 0.000 description 4
- 108050008953 Melanoma-associated antigen Proteins 0.000 description 4
- 108010072866 Prostate-Specific Antigen Proteins 0.000 description 4
- 102100038358 Prostate-specific antigen Human genes 0.000 description 4
- IQFYYKKMVGJFEH-XLPZGREQSA-N Thymidine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 IQFYYKKMVGJFEH-XLPZGREQSA-N 0.000 description 4
- 102100036856 Tumor necrosis factor receptor superfamily member 9 Human genes 0.000 description 4
- ISAKRJDGNUQOIC-UHFFFAOYSA-N Uracil Chemical compound O=C1C=CNC(=O)N1 ISAKRJDGNUQOIC-UHFFFAOYSA-N 0.000 description 4
- 102100026497 Zinc finger protein 654 Human genes 0.000 description 4
- 108010026331 alpha-Fetoproteins Proteins 0.000 description 4
- 230000008901 benefit Effects 0.000 description 4
- 239000003795 chemical substances by application Substances 0.000 description 4
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Chemical compound NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 4
- 238000013461 design Methods 0.000 description 4
- 229940126534 drug product Drugs 0.000 description 4
- 238000013401 experimental design Methods 0.000 description 4
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 4
- 239000000833 heterodimer Substances 0.000 description 4
- 238000009169 immunotherapy Methods 0.000 description 4
- 208000015181 infectious disease Diseases 0.000 description 4
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 4
- 108020001756 ligand binding domains Proteins 0.000 description 4
- 239000012528 membrane Substances 0.000 description 4
- 239000000178 monomer Substances 0.000 description 4
- 239000013642 negative control Substances 0.000 description 4
- 239000000825 pharmaceutical preparation Substances 0.000 description 4
- 102100030886 Complement receptor type 1 Human genes 0.000 description 3
- 108090000331 Firefly luciferases Proteins 0.000 description 3
- 101000727061 Homo sapiens Complement receptor type 1 Proteins 0.000 description 3
- 101001137987 Homo sapiens Lymphocyte activation gene 3 protein Proteins 0.000 description 3
- 108010021625 Immunoglobulin Fragments Proteins 0.000 description 3
- 102000008394 Immunoglobulin Fragments Human genes 0.000 description 3
- 208000031671 Large B-Cell Diffuse Lymphoma Diseases 0.000 description 3
- 206010061309 Neoplasm progression Diseases 0.000 description 3
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 3
- 102100040247 Tumor necrosis factor Human genes 0.000 description 3
- 230000003213 activating effect Effects 0.000 description 3
- 230000000735 allogeneic effect Effects 0.000 description 3
- 238000013459 approach Methods 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 230000001413 cellular effect Effects 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 206010012818 diffuse large B-cell lymphoma Diseases 0.000 description 3
- 231100000673 dose–response relationship Toxicity 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000000684 flow cytometry Methods 0.000 description 3
- 239000003102 growth factor Substances 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 230000028709 inflammatory response Effects 0.000 description 3
- 230000003993 interaction Effects 0.000 description 3
- 238000007918 intramuscular administration Methods 0.000 description 3
- 238000004020 luminiscence type Methods 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 230000007246 mechanism Effects 0.000 description 3
- 201000000050 myeloid neoplasm Diseases 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 210000002966 serum Anatomy 0.000 description 3
- 238000007920 subcutaneous administration Methods 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 238000002560 therapeutic procedure Methods 0.000 description 3
- 230000005751 tumor progression Effects 0.000 description 3
- LRFVTYWOQMYALW-UHFFFAOYSA-N 9H-xanthine Chemical compound O=C1NC(=O)NC2=C1NC=N2 LRFVTYWOQMYALW-UHFFFAOYSA-N 0.000 description 2
- 229930024421 Adenine Natural products 0.000 description 2
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 2
- 102100036475 Alanine aminotransferase 1 Human genes 0.000 description 2
- 108010082126 Alanine transaminase Proteins 0.000 description 2
- 101710145634 Antigen 1 Proteins 0.000 description 2
- 108010031480 Artificial Receptors Proteins 0.000 description 2
- 108010003415 Aspartate Aminotransferases Proteins 0.000 description 2
- 102000004625 Aspartate Aminotransferases Human genes 0.000 description 2
- 102100029822 B- and T-lymphocyte attenuator Human genes 0.000 description 2
- 102100038080 B-cell receptor CD22 Human genes 0.000 description 2
- 102100021663 Baculoviral IAP repeat-containing protein 5 Human genes 0.000 description 2
- DWRXFEITVBNRMK-UHFFFAOYSA-N Beta-D-1-Arabinofuranosylthymine Natural products O=C1NC(=O)C(C)=CN1C1C(O)C(O)C(CO)O1 DWRXFEITVBNRMK-UHFFFAOYSA-N 0.000 description 2
- 102100024263 CD160 antigen Human genes 0.000 description 2
- 102100027207 CD27 antigen Human genes 0.000 description 2
- 101150013553 CD40 gene Proteins 0.000 description 2
- 102100035793 CD83 antigen Human genes 0.000 description 2
- 108010051152 Carboxylesterase Proteins 0.000 description 2
- 102000013392 Carboxylesterase Human genes 0.000 description 2
- 102100025064 Cellular tumor antigen p53 Human genes 0.000 description 2
- 102000057710 Coatomer Human genes 0.000 description 2
- 108091026890 Coding region Proteins 0.000 description 2
- 241000702421 Dependoparvovirus Species 0.000 description 2
- 101150029707 ERBB2 gene Proteins 0.000 description 2
- 208000032612 Glial tumor Diseases 0.000 description 2
- 206010018338 Glioma Diseases 0.000 description 2
- 102100041003 Glutamate carboxypeptidase 2 Human genes 0.000 description 2
- 102100034458 Hepatitis A virus cellular receptor 2 Human genes 0.000 description 2
- 241000282412 Homo Species 0.000 description 2
- 101000864344 Homo sapiens B- and T-lymphocyte attenuator Proteins 0.000 description 2
- 101000884305 Homo sapiens B-cell receptor CD22 Proteins 0.000 description 2
- 101000761938 Homo sapiens CD160 antigen Proteins 0.000 description 2
- 101000914511 Homo sapiens CD27 antigen Proteins 0.000 description 2
- 101000946856 Homo sapiens CD83 antigen Proteins 0.000 description 2
- 101000721661 Homo sapiens Cellular tumor antigen p53 Proteins 0.000 description 2
- 101000892862 Homo sapiens Glutamate carboxypeptidase 2 Proteins 0.000 description 2
- 101001068133 Homo sapiens Hepatitis A virus cellular receptor 2 Proteins 0.000 description 2
- 101000777628 Homo sapiens Leukocyte antigen CD37 Proteins 0.000 description 2
- 101000623901 Homo sapiens Mucin-16 Proteins 0.000 description 2
- 101001109503 Homo sapiens NKG2-C type II integral membrane protein Proteins 0.000 description 2
- 101000831007 Homo sapiens T-cell immunoreceptor with Ig and ITIM domains Proteins 0.000 description 2
- 101000655352 Homo sapiens Telomerase reverse transcriptase Proteins 0.000 description 2
- 101000851376 Homo sapiens Tumor necrosis factor receptor superfamily member 8 Proteins 0.000 description 2
- 241000598436 Human T-cell lymphotropic virus Species 0.000 description 2
- 108010031794 IGF Type 1 Receptor Proteins 0.000 description 2
- 102000004877 Insulin Human genes 0.000 description 2
- 108090001061 Insulin Proteins 0.000 description 2
- 102000048143 Insulin-Like Growth Factor II Human genes 0.000 description 2
- 108090001117 Insulin-Like Growth Factor II Proteins 0.000 description 2
- 102100039688 Insulin-like growth factor 1 receptor Human genes 0.000 description 2
- 102100034872 Kallikrein-4 Human genes 0.000 description 2
- 101150030213 Lag3 gene Proteins 0.000 description 2
- 108010028275 Leukocyte Elastase Proteins 0.000 description 2
- 102100031586 Leukocyte antigen CD37 Human genes 0.000 description 2
- 108010046938 Macrophage Colony-Stimulating Factor Proteins 0.000 description 2
- 102100028123 Macrophage colony-stimulating factor 1 Human genes 0.000 description 2
- 102000003735 Mesothelin Human genes 0.000 description 2
- 108090000015 Mesothelin Proteins 0.000 description 2
- 241000204031 Mycoplasma Species 0.000 description 2
- 102100022683 NKG2-C type II integral membrane protein Human genes 0.000 description 2
- 102100033174 Neutrophil elastase Human genes 0.000 description 2
- 206010034016 Paronychia Diseases 0.000 description 2
- 102100034937 Poly(A) RNA polymerase, mitochondrial Human genes 0.000 description 2
- 206010060862 Prostate cancer Diseases 0.000 description 2
- 102000001253 Protein Kinase Human genes 0.000 description 2
- 108020004511 Recombinant DNA Proteins 0.000 description 2
- 241000700584 Simplexvirus Species 0.000 description 2
- 102100037253 Solute carrier family 45 member 3 Human genes 0.000 description 2
- 108010002687 Survivin Proteins 0.000 description 2
- 102100024834 T-cell immunoreceptor with Ig and ITIM domains Human genes 0.000 description 2
- 108700012920 TNF Proteins 0.000 description 2
- 108010017842 Telomerase Proteins 0.000 description 2
- 101710199837 Terminal uridylyltransferase 1 Proteins 0.000 description 2
- 102000009843 Thyroglobulin Human genes 0.000 description 2
- 108010034949 Thyroglobulin Proteins 0.000 description 2
- 101000642183 Trypanosoma brucei brucei Terminal uridylyltransferase 2 Proteins 0.000 description 2
- 102100040245 Tumor necrosis factor receptor superfamily member 5 Human genes 0.000 description 2
- 102100036857 Tumor necrosis factor receptor superfamily member 8 Human genes 0.000 description 2
- 102000003425 Tyrosinase Human genes 0.000 description 2
- 108060008724 Tyrosinase Proteins 0.000 description 2
- SAZUGELZHZOXHB-UHFFFAOYSA-N acecarbromal Chemical compound CCC(Br)(CC)C(=O)NC(=O)NC(C)=O SAZUGELZHZOXHB-UHFFFAOYSA-N 0.000 description 2
- 229960000643 adenine Drugs 0.000 description 2
- 239000002671 adjuvant Substances 0.000 description 2
- 239000000443 aerosol Substances 0.000 description 2
- 230000000890 antigenic effect Effects 0.000 description 2
- 239000003963 antioxidant agent Substances 0.000 description 2
- IQFYYKKMVGJFEH-UHFFFAOYSA-N beta-L-thymidine Natural products O=C1NC(=O)C(C)=CN1C1OC(CO)C(O)C1 IQFYYKKMVGJFEH-UHFFFAOYSA-N 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 150000001720 carbohydrates Chemical class 0.000 description 2
- 235000014633 carbohydrates Nutrition 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 230000032823 cell division Effects 0.000 description 2
- 238000002659 cell therapy Methods 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 238000012512 characterization method Methods 0.000 description 2
- 239000012707 chemical precursor Substances 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000002591 computed tomography Methods 0.000 description 2
- 238000011443 conventional therapy Methods 0.000 description 2
- 108091008034 costimulatory receptors Proteins 0.000 description 2
- 229940104302 cytosine Drugs 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 230000007850 degeneration Effects 0.000 description 2
- 230000003111 delayed effect Effects 0.000 description 2
- 229910003460 diamond Inorganic materials 0.000 description 2
- 239000010432 diamond Substances 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- 201000003444 follicular lymphoma Diseases 0.000 description 2
- 239000012595 freezing medium Substances 0.000 description 2
- 235000011187 glycerol Nutrition 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 208000021173 high grade B-cell lymphoma Diseases 0.000 description 2
- 229940084986 human chorionic gonadotropin Drugs 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- 230000036039 immunity Effects 0.000 description 2
- 238000013339 in-process testing Methods 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 230000000977 initiatory effect Effects 0.000 description 2
- 229940125396 insulin Drugs 0.000 description 2
- 230000000968 intestinal effect Effects 0.000 description 2
- 230000003834 intracellular effect Effects 0.000 description 2
- 108010024383 kallikrein 4 Proteins 0.000 description 2
- 238000011005 laboratory method Methods 0.000 description 2
- 230000000670 limiting effect Effects 0.000 description 2
- 210000001165 lymph node Anatomy 0.000 description 2
- 231100000682 maximum tolerated dose Toxicity 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- -1 mut hsp70-2 Proteins 0.000 description 2
- 108700043045 nanoluc Proteins 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- 229920001481 poly(stearyl methacrylate) Polymers 0.000 description 2
- 108040000983 polyphosphate:AMP phosphotransferase activity proteins Proteins 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 201000006037 primary mediastinal B-cell lymphoma Diseases 0.000 description 2
- 201000001514 prostate carcinoma Diseases 0.000 description 2
- 108010079891 prostein Proteins 0.000 description 2
- 108060006633 protein kinase Proteins 0.000 description 2
- 230000009257 reactivity Effects 0.000 description 2
- 230000006798 recombination Effects 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- 238000012429 release testing Methods 0.000 description 2
- 230000000717 retained effect Effects 0.000 description 2
- 230000001177 retroviral effect Effects 0.000 description 2
- 231100000004 severe toxicity Toxicity 0.000 description 2
- 210000000130 stem cell Anatomy 0.000 description 2
- 239000007929 subcutaneous injection Substances 0.000 description 2
- 229940104230 thymidine Drugs 0.000 description 2
- 210000001541 thymus gland Anatomy 0.000 description 2
- 229960002175 thyroglobulin Drugs 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 241000701161 unidentified adenovirus Species 0.000 description 2
- 241001430294 unidentified retrovirus Species 0.000 description 2
- 229940035893 uracil Drugs 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- LDGWQMRUWMSZIU-LQDDAWAPSA-M 2,3-bis[(z)-octadec-9-enoxy]propyl-trimethylazanium;chloride Chemical compound [Cl-].CCCCCCCC\C=C/CCCCCCCCOCC(C[N+](C)(C)C)OCCCCCCCC\C=C/CCCCCCCC LDGWQMRUWMSZIU-LQDDAWAPSA-M 0.000 description 1
- 241001664176 Alpharetrovirus Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 241000282472 Canis lupus familiaris Species 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 201000009030 Carcinoma Diseases 0.000 description 1
- 102000014914 Carrier Proteins Human genes 0.000 description 1
- 241000700198 Cavia Species 0.000 description 1
- 206010057248 Cell death Diseases 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 108091007741 Chimeric antigen receptor T cells Proteins 0.000 description 1
- CMSMOCZEIVJLDB-UHFFFAOYSA-N Cyclophosphamide Chemical compound ClCCN(CCCl)P1(=O)NCCCO1 CMSMOCZEIVJLDB-UHFFFAOYSA-N 0.000 description 1
- 108010025905 Cystine-Knot Miniproteins Proteins 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- 102000053602 DNA Human genes 0.000 description 1
- 229920002307 Dextran Polymers 0.000 description 1
- 101710140859 E3 ubiquitin ligase TRAF3IP2 Proteins 0.000 description 1
- 102100026620 E3 ubiquitin ligase TRAF3IP2 Human genes 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 241000792859 Enema Species 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000283086 Equidae Species 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 102000000802 Galectin 3 Human genes 0.000 description 1
- 108010001517 Galectin 3 Proteins 0.000 description 1
- 241001663880 Gammaretrovirus Species 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 208000009329 Graft vs Host Disease Diseases 0.000 description 1
- 208000002250 Hematologic Neoplasms Diseases 0.000 description 1
- 101001030211 Homo sapiens Myc proto-oncogene protein Proteins 0.000 description 1
- 101000801254 Homo sapiens Tumor necrosis factor receptor superfamily member 16 Proteins 0.000 description 1
- 241000713673 Human foamy virus Species 0.000 description 1
- 241000701806 Human papillomavirus Species 0.000 description 1
- 241001135958 Human type D retrovirus Species 0.000 description 1
- 206010020751 Hypersensitivity Diseases 0.000 description 1
- UGQMRVRMYYASKQ-UHFFFAOYSA-N Hypoxanthine nucleoside Natural products OC1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 UGQMRVRMYYASKQ-UHFFFAOYSA-N 0.000 description 1
- 102000037978 Immune checkpoint receptors Human genes 0.000 description 1
- 108091008028 Immune checkpoint receptors Proteins 0.000 description 1
- 206010061598 Immunodeficiency Diseases 0.000 description 1
- 208000029462 Immunodeficiency disease Diseases 0.000 description 1
- 102000016844 Immunoglobulin-like domains Human genes 0.000 description 1
- 108050006430 Immunoglobulin-like domains Proteins 0.000 description 1
- 108090000174 Interleukin-10 Proteins 0.000 description 1
- 102000003814 Interleukin-10 Human genes 0.000 description 1
- 108010002586 Interleukin-7 Proteins 0.000 description 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 1
- 108090001090 Lectins Proteins 0.000 description 1
- 102000004856 Lectins Human genes 0.000 description 1
- 108060001084 Luciferase Proteins 0.000 description 1
- 239000005089 Luciferase Substances 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 206010027476 Metastases Diseases 0.000 description 1
- 101100341510 Mus musculus Itgal gene Proteins 0.000 description 1
- 238000000636 Northern blotting Methods 0.000 description 1
- 108700026244 Open Reading Frames Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 102000010292 Peptide Elongation Factor 1 Human genes 0.000 description 1
- 108010077524 Peptide Elongation Factor 1 Proteins 0.000 description 1
- 102000016611 Proteoglycans Human genes 0.000 description 1
- 108010067787 Proteoglycans Proteins 0.000 description 1
- 102000052575 Proto-Oncogene Human genes 0.000 description 1
- 108700020978 Proto-Oncogene Proteins 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 description 1
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 description 1
- 206010039491 Sarcoma Diseases 0.000 description 1
- 108010003723 Single-Domain Antibodies Proteins 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 238000002105 Southern blotting Methods 0.000 description 1
- 241000713675 Spumavirus Species 0.000 description 1
- 101000677856 Stenotrophomonas maltophilia (strain K279a) Actin-binding protein Smlt3054 Proteins 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 230000024932 T cell mediated immunity Effects 0.000 description 1
- 230000006052 T cell proliferation Effects 0.000 description 1
- 108060008683 Tumor Necrosis Factor Receptor Proteins 0.000 description 1
- 101710187888 Tumor necrosis factor receptor superfamily member 16 Proteins 0.000 description 1
- 241000700618 Vaccinia virus Species 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 210000004504 adult stem cell Anatomy 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 230000002776 aggregation Effects 0.000 description 1
- 238000004220 aggregation Methods 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 208000026935 allergic disease Diseases 0.000 description 1
- 230000007815 allergy Effects 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- WNROFYMDJYEPJX-UHFFFAOYSA-K aluminium hydroxide Chemical compound [OH-].[OH-].[OH-].[Al+3] WNROFYMDJYEPJX-UHFFFAOYSA-K 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000004873 anchoring Methods 0.000 description 1
- 230000000692 anti-sense effect Effects 0.000 description 1
- 230000006023 anti-tumor response Effects 0.000 description 1
- 238000011319 anticancer therapy Methods 0.000 description 1
- 210000000612 antigen-presenting cell Anatomy 0.000 description 1
- 230000005975 antitumor immune response Effects 0.000 description 1
- 238000002617 apheresis Methods 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 230000005784 autoimmunity Effects 0.000 description 1
- 230000006399 behavior Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 108091008324 binding proteins Proteins 0.000 description 1
- 238000005415 bioluminescence Methods 0.000 description 1
- 230000029918 bioluminescence Effects 0.000 description 1
- 201000000053 blastoma Diseases 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 239000012888 bovine serum Substances 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 239000007975 buffered saline Substances 0.000 description 1
- 239000008366 buffered solution Substances 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000024245 cell differentiation Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 239000002738 chelating agent Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000006690 co-activation Effects 0.000 description 1
- 238000001553 co-assembly Methods 0.000 description 1
- 238000002648 combination therapy Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 238000004590 computer program Methods 0.000 description 1
- 230000006552 constitutive activation Effects 0.000 description 1
- 239000000356 contaminant Substances 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 230000000139 costimulatory effect Effects 0.000 description 1
- 229960004397 cyclophosphamide Drugs 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 231100000433 cytotoxic Toxicity 0.000 description 1
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 1
- 230000001472 cytotoxic effect Effects 0.000 description 1
- 230000007402 cytotoxic response Effects 0.000 description 1
- 230000002950 deficient Effects 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 239000000539 dimer Substances 0.000 description 1
- 230000000447 dimerizing effect Effects 0.000 description 1
- 230000003467 diminishing effect Effects 0.000 description 1
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 1
- 208000035475 disorder Diseases 0.000 description 1
- 125000002228 disulfide group Chemical group 0.000 description 1
- 230000003828 downregulation Effects 0.000 description 1
- 239000006196 drop Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000003221 ear drop Substances 0.000 description 1
- 229940047652 ear drops Drugs 0.000 description 1
- 230000002500 effect on skin Effects 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 201000008184 embryoma Diseases 0.000 description 1
- 210000001671 embryonic stem cell Anatomy 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 229940095399 enema Drugs 0.000 description 1
- 239000007920 enema Substances 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 239000013604 expression vector Substances 0.000 description 1
- 238000013265 extended release Methods 0.000 description 1
- 239000003889 eye drop Substances 0.000 description 1
- 229940012356 eye drops Drugs 0.000 description 1
- 210000004700 fetal blood Anatomy 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 229960000390 fludarabine Drugs 0.000 description 1
- GIUYCYHIANZCFB-FJFJXFQQSA-N fludarabine phosphate Chemical compound C1=NC=2C(N)=NC(F)=NC=2N1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)[C@@H]1O GIUYCYHIANZCFB-FJFJXFQQSA-N 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 210000004475 gamma-delta t lymphocyte Anatomy 0.000 description 1
- 238000012239 gene modification Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 230000005017 genetic modification Effects 0.000 description 1
- 235000013617 genetically modified food Nutrition 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 1
- 208000024908 graft versus host disease Diseases 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 230000003370 grooming effect Effects 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- 210000002443 helper t lymphocyte Anatomy 0.000 description 1
- 201000005787 hematologic cancer Diseases 0.000 description 1
- 230000002489 hematologic effect Effects 0.000 description 1
- 208000024200 hematopoietic and lymphoid system neoplasm Diseases 0.000 description 1
- 102000053563 human MYC Human genes 0.000 description 1
- 102000052073 human NGFR Human genes 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
- 102000027596 immune receptors Human genes 0.000 description 1
- 108091008915 immune receptors Proteins 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000007813 immunodeficiency Effects 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 210000004263 induced pluripotent stem cell Anatomy 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000011221 initial treatment Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 238000001361 intraarterial administration Methods 0.000 description 1
- 238000000185 intracerebroventricular administration Methods 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- 239000007928 intraperitoneal injection Substances 0.000 description 1
- 238000007913 intrathecal administration Methods 0.000 description 1
- 238000010253 intravenous injection Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 150000002605 large molecules Chemical class 0.000 description 1
- 230000002045 lasting effect Effects 0.000 description 1
- 239000002523 lectin Substances 0.000 description 1
- 238000009092 lines of therapy Methods 0.000 description 1
- 238000001638 lipofection Methods 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 239000012669 liquid formulation Substances 0.000 description 1
- 230000004807 localization Effects 0.000 description 1
- 230000033001 locomotion Effects 0.000 description 1
- 231100001252 long-term toxicity Toxicity 0.000 description 1
- 238000011469 lymphodepleting chemotherapy Methods 0.000 description 1
- 239000008176 lyophilized powder Substances 0.000 description 1
- 108010026228 mRNA guanylyltransferase Proteins 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 210000002284 membrane microdomain Anatomy 0.000 description 1
- 210000003071 memory t lymphocyte Anatomy 0.000 description 1
- 230000009401 metastasis Effects 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 238000010369 molecular cloning Methods 0.000 description 1
- 230000004899 motility Effects 0.000 description 1
- 238000010172 mouse model Methods 0.000 description 1
- 210000000581 natural killer T-cell Anatomy 0.000 description 1
- 210000000822 natural killer cell Anatomy 0.000 description 1
- 230000001613 neoplastic effect Effects 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 239000002777 nucleoside Substances 0.000 description 1
- 150000003833 nucleoside derivatives Chemical class 0.000 description 1
- 229950002610 otelixizumab Drugs 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 239000000816 peptidomimetic Substances 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 239000002953 phosphate buffered saline Substances 0.000 description 1
- 229920005862 polyol Polymers 0.000 description 1
- 150000003077 polyols Chemical class 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 238000002600 positron emission tomography Methods 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000012910 preclinical development Methods 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 208000037920 primary disease Diseases 0.000 description 1
- 208000037821 progressive disease Diseases 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 230000000069 prophylactic effect Effects 0.000 description 1
- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 230000002797 proteolythic effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 210000003289 regulatory T cell Anatomy 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 238000003757 reverse transcription PCR Methods 0.000 description 1
- 229920002477 rna polymer Polymers 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 230000033237 signal complex assembly Effects 0.000 description 1
- 150000003384 small molecules Chemical class 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 230000009870 specific binding Effects 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 238000010254 subcutaneous injection Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 230000003319 supportive effect Effects 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 230000004654 survival pathway Effects 0.000 description 1
- 206010042863 synovial sarcoma Diseases 0.000 description 1
- 238000010189 synthetic method Methods 0.000 description 1
- 239000003826 tablet Substances 0.000 description 1
- 229950010127 teplizumab Drugs 0.000 description 1
- 238000011287 therapeutic dose Methods 0.000 description 1
- 230000000699 topical effect Effects 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 230000002463 transducing effect Effects 0.000 description 1
- 230000009261 transgenic effect Effects 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- 230000005909 tumor killing Effects 0.000 description 1
- 102000003298 tumor necrosis factor receptor Human genes 0.000 description 1
- 235000015112 vegetable and seed oil Nutrition 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 229950004393 visilizumab Drugs 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
- 229940075420 xanthine Drugs 0.000 description 1
- 238000012447 xenograft mouse model Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
- C07K14/70596—Molecules with a "CD"-designation not provided for elsewhere
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0634—Cells from the blood or the immune system
- C12N5/0636—T lymphocytes
-
- 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
- A61K39/464—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
- 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
- A61K39/464416—Receptors for cytokines
- A61K39/464417—Receptors for tumor necrosis factors [TNF], e.g. lymphotoxin receptor [LTR], CD30
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
- C07K14/70503—Immunoglobulin superfamily
- C07K14/7051—T-cell receptor (TcR)-CD3 complex
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
- C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
- C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
- C07K16/2809—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily against the T-cell receptor (TcR)-CD3 complex
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
- C07K16/2878—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/32—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against translation products of oncogenes
-
- 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
- A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
-
- 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
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/20—Immunoglobulins specific features characterized by taxonomic origin
- C07K2317/24—Immunoglobulins specific features characterized by taxonomic origin containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
- C07K2317/31—Immunoglobulins specific features characterized by aspects of specificity or valency multispecific
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/60—Immunoglobulins specific features characterized by non-natural combinations of immunoglobulin fragments
- C07K2317/62—Immunoglobulins specific features characterized by non-natural combinations of immunoglobulin fragments comprising only variable region components
- C07K2317/622—Single chain antibody (scFv)
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
- C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
- C07K2319/03—Fusion polypeptide containing a localisation/targetting motif containing a transmembrane segment
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
- C07K2319/33—Fusion polypeptide fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
Definitions
- nucleic acid sequences encoding a CD 19 Trifunctional T cell-antigen coupler comprises: (a) a first polynucleotide encoding a ligand that selectively binds a CD 19 antigen.
- the nucleic acid sequence encoding a CD 19 Trifunctional T cell-antigen coupler comprises: (b) a second polynucleotide encoding a UCHT1 ligand that binds CD3.
- the nucleic acid sequence encoding a CD 19 Trifunctional T cell-antigen coupler comprise: (c) a third polynucleotide encoding a TCR signaling domain polypeptide comprising a cytosolic domain and a transmembrane domain.
- the components encoded by the first, second, and/or third polynucleotides are connected in any suitable manner, such as in any suitable order and/or comprising any suitable linker(s).
- the components encoded by (a), components encoded by (b), and components encoded by (c) are fused directly to each other, or joined by at least one linker.
- the ligand that selectively binds the CD 19 antigen is a single chain variable fragment (scFv).
- the ligand that selectively binds the CD 19 antigen comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 36.
- the UCHT1 ligand is a single chain antibody.
- the UCHT1 ligand comprises a Y182T mutation (SEQ ID NO: 72).
- the UCHT1 ligand is a humanized variant of UCHT1 (huUCHTl) ligand (SEQ ID NO: 44).
- the UCHT1 ligand is a humanized variant of UCHT1 comprising a Y177T mutation (huUCHTl (Y177T)) (SEQ ID NO: 46).
- the UCHT1 ligand comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 14, SEQ ID NO: 72, SEQ ID NO: 44, or SEQ ID NO: 46.
- the cytosolic domain is a CD4 cytosolic domain and the transmembrane domain is a CD4 transmembrane domain.
- the third polynucleotide encodes a polypeptide comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 18.
- the component encoded by (a) and the component encoded by (c) are fused to the component encoded by (b).
- the component encoded by (b) and the component encoded by (c) are fused to the component encoded by (a).
- at least one linker joins the component encoded by (a) to the component encoded by (b).
- the at least one linker is a G 4 S flexible linker (SEQ ID NO: 73), a large protein domain, a long helix structure, or a short helix structure.
- the at least one linker comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 12 (G 4 S flexible linker ("G 4 S" disclosed as SEQ ID NO: 73)), SEQ ID NO: 32 (large protein domain), SEQ ID NO: 30 (long helix structure), or SEQ ID NO: 28 (short helix structure).
- the CD3 is of a TCR complex on a cell expressing the second polynucleotide. In some embodiments, the binding of the CD3 induces activation of a cell expressing the second polynucleotide.
- the CD19-TAC comprises a nucleic acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 64.
- the nucleic acid sequence does not encode a co-stimulatory domain.
- the nucleic acid sequence does not encode an activation domain.
- vector constructs comprising: (a) a nucleic acid sequence disclosed herein (e.g., a nucleic acid sequence encoding a CD19-TAC); and (b) a promoter functional in a mammalian cell.
- T cells comprising a nucleic acid sequence disclosed herein (e.g., a nucleic acid sequence encoding a CD19-TAC).
- compositions comprising the T cell disclosed herein, and a pharmaceutically acceptable excipient.
- the cancer is a B cell malignancy.
- the cancer is B cell lymphoma, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), or Non-Hodgkins Lymphoma.
- the pharmaceutical composition is administered transarterially, subcutaneously, intradermally, intratumorally, intranodally, intramedullary, intramuscularly, intravenously or intraperitoneally.
- nucleic acid sequences encoding a Trifunctional T cell-antigen coupler comprising: (a) a first polynucleotide encoding a target-specific ligand; (b) a second polynucleotide encoding a ligand that binds a protein associated with a TCR complex; and (c) a third polynucleotide encoding a T cell receptor signaling domain polypeptide; wherein the ligand that binds the protein associated with the TCR complex is selected from OKT3, F6A or L2K.
- Tri-TAC Trifunctional T cell-antigen coupler
- component encoded by (a), component encoded by (b), and component encoded by (c) are fused directly to each other, or joined by at least one linker. In some embodiments, the component encoded by (a) and the component encoded by (b) are directly fused and joined to the component encoded by (c) by a linker. In some embodiments, the component encoded by (b) and the component encoded by (c) are directly fused and joined to the component encoded by (a) by a linker. In some
- the at least one linker is a G 4 S flexible linker (SEQ ID NO: 73), a large protein domain, a long helix structure, or a short helix structure.
- the at least one linker has an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 12 (G 4 S flexible linker ("G 4 S" disclosed as SEQ ID NO: 73)), SEQ ID NO: 32 (large protein domain), SEQ ID NO: 30 (long helix structure), or SEQ ID NO: 28 (short helix structure).
- the ligand that binds the protein associated with the TCR complex is OKT3.
- the ligand that binds a protein associated with the TCR complex comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 22.
- the ligand that binds the protein associated with the TCR complex is F6A.
- the ligand that binds the protein associated with the TCR complex comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 24.
- the ligand that binds the protein associated with the TCR complex is L2K.
- the ligand that binds the protein associated with the TCR complex comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 26.
- the protein associated with the TCR complex is CD3.
- the target-specific ligand selectively binds a tumor antigen.
- the target-specific ligand is a designed ankyrin repeat (DARPin) polypeptide, or a single chain variable fragment (scFv).
- DARPin ankyrin repeat
- scFv single chain variable fragment
- the target-specific ligand selectively binds a CD 19 antigen, a HER2 antigen, or a BCMA antigen.
- the target-specific ligand selectively binds a HER-2 antigen comprises an antigen binding domain of an antibody selected from Trastuzumab, Pertuzumab, Lapatinib, Neratinib, Ado-trastuzmab Emtansine, Gancotamab, Margetuximab, Timigutuzumab, and Ertumaxomab.
- the target-specific ligand selectively binds a BCMA antigen comprises an antigen binding domain of an antibody selected from Belantamab mafodotin, and GSK2857916.
- the target-specific ligand comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 36, SEQ ID NO: 8 or SEQ ID NO: 34.
- the T cell receptor signaling domain polypeptide comprises a cytosolic domain and a transmembrane domain.
- the cytosolic domain is a CD4 cytosolic domain and the transmembrane domain is a CD4 transmembrane domain, or wherein the cytosolic domain is a CD8 cytosolic domain and the transmembrane domain is a CD8 transmembrane domain.
- the nucleic acid sequences further comprise a leader sequence.
- the leader sequence comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 6, SEQ ID NO: 48, or SEQ ID NO: 50.
- the CD3 is of a TCR complex on a cell expressing the second polynucleotide. In some embodiments, the binding of the CD3 induces activation of a cell expressing the second polynucleotide.
- the Tri-TAC comprises a nucleic acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 75, SEQ ID NO: 55, SEQ ID NO: 57, SEQ ID NO: 59, or SEQ ID NO: 61.
- the Tri-TAC comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 64, SEQ ID NO: 66, SEQ ID NO: 68, SEQ ID NO: 76, SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 60, or SEQ ID NO: 62.
- the nucleic acid sequence does not encode a co- stimulatory domain. In some embodiments, the nucleic acid sequence does not encode an activation domain.
- nucleic acid sequences encoding a Trifunctional T cell-antigen coupler comprising: (a) a first polynucleotide encoding a target-specific ligand; (b) a second polynucleotide encoding a ligand that binds a protein associated with a TCR complex; and (c) a third polynucleotide encoding a T cell receptor signaling domain polypeptide; wherein the nucleic acid sequence further comprises a leader sequence, and wherein component encoded by (a), component encoded by (b), and component encoded by (c) are fused directly to each other, or joined by at least one linker.
- the target-specific ligand selectively binds a tumor antigen.
- the target-specific ligand is a designed ankyrin repeat (DARPin) polypeptide, or a single chain variable fragment (scFv).
- DARPin ankyrin repeat
- scFv single chain variable fragment
- the target-specific ligand selectively binds a CD 19 antigen, a HER2 antigen, or a BCMA antigen.
- the target-specific ligand selectively binds a HER-2 antigen comprises an antigen binding domain of an antibody selected from Trastuzumab, Pertuzumab, Lapatinib, Neratinib, Ado- trastuzmab Emtansine, Gancotamab, Margetuximab, Timigutuzumab, and Ertumaxomab.
- the target-specific ligand selectively binds a BCMA antigen comprises an antigen binding domain of an antibody selected from Belantamab mafodotin, and GSK2857916.
- the target-specific ligand comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 36, SEQ ID NO: 8, SEQ ID NO: 34, SEQ ID NO: 52, or SEQ ID NO: 54.
- the ligand that binds the protein associated with the TCR complex is selected from UCHT1, UCHT1 (Y182T), huUCHTl, huUCHTl (Y177T), OKT3, F6A, or L2K.
- the ligand that binds a protein associated with the TCR complex has an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 14, SEQ ID NO: 72, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 22, SEQ ID NO: 24, or SEQ ID NO: 26.
- the protein associated with the TCR complex is CD3.
- the T cell receptor signaling domain polypeptide comprises a cytosolic domain and a transmembrane domain.
- the cytosolic domain is a CD4 cytosolic domain and the transmembrane domain is a CD4 transmembrane domain, or wherein the cytosolic domain is a CD8 cytosolic domain and the transmembrane domain is a CD8 transmembrane domain.
- the leader sequence comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 6, SEQ ID NO: 48, or SEQ ID NO: 50.
- the component encoded by (a) and the component encoded by (b) are directly fused and joined to the component encoded by (c) by a linker.
- the component encoded by (b) and the component encoded by (c) are directly fused and joined to the component encoded by (a) by a linker.
- the at least one linker is a G 4 S flexible linker (SEQ ID NO: 73), a large protein domain, a long helix structure, or a short helix structure.
- the at least one linker has an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 12 (G 4 S flexible linker ("G 4 S" disclosed as SEQ ID NO: 73)), SEQ ID NO: 32 (large protein domain), SEQ ID NO: 30 (long helix structure), or SEQ ID NO: 28 (short helix structure).
- the CD3 is of a TCR complex on a cell expressing the second polynucleotide. In some embodiments, the binding of the CD3 induces activation of a cell expressing the second polynucleotide.
- the Tri-TAC comprises a nucleic acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 75, SEQ ID NO: 55, SEQ ID NO: 57, SEQ ID NO: 59, or SEQ ID NO: 61.
- the Tri-TAC comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 64, SEQ ID NO: 66, SEQ ID NO: 68, SEQ ID NO: 76, SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 60, or SEQ ID NO: 62.
- the nucleic acid sequence does not encode a co- stimulatory domain. In some embodiments, the nucleic acid sequence does not encode an activation domain.
- polypeptides encoded by the nucleic acid sequence disclosed herein are polypeptides encoded by the nucleic acid sequence disclosed herein.
- vector constructs comprising: (a) a nucleic acid sequence disclosed herein; and (b) a promoter functional in a mammalian cell.
- T cells comprising the nucleic acid sequence disclosed herein.
- compositions comprising the T cell disclosed herein, and a pharmaceutically acceptable excipient.
- the cancer is a solid cancer or a liquid cancer.
- the cancer is a lung cancer, a breast cancer, multiple myeloma, glioblastoma, gastric cancer, ovarian cancer, stomach cancer, colorectal cancer, urothelial cancer, endometrial cancer, or a colon cancer.
- the cancer comprises a CD 19 expressing cancer cell.
- the cancer is a B cell malignancy.
- the cancer is B cell lymphoma, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), or Non-Hodgkins Lymphoma.
- the cancer comprises a HER-2 expressing cancer cell.
- the cancer is breast cancer, bladder cancer, pancreatic cancer, ovarian cancer, or stomach cancer.
- the cancer comprises a BCMA expressing cancer cell.
- the cancer is leukemia, lymphoma, or multiple myeloma.
- the pharmaceutical composition is administered to the individual transarterially, subcutaneously, intradermally, intratumorally, intranodally, intramedullary, intramuscularly, intravenously or intraperitoneally.
- the pharmaceutical composition is in a unit dose form.
- the pharmaceutical composition comprises about 0.5 - 2 x 10 9 T cells.
- the pharmaceutical composition is administered daily, weekly, bi-weekly, monthly, bi-month or yearly.
- nucleic acid sequences encoding a CD 19 Trifunctional T cell-antigen coupler comprising: (a) a first polynucleotide encoding a ligand that selectively binds a CD 19 antigen; (b) a second polynucleotide encoding a humanized variant of a UCHT1 (huUCHTl) ligand comprising a Y177T mutation (huUCHTl (Y177T)) that binds CD3; and (c) a third polynucleotide encoding a polypeptide comprising a CD4 cytosolic domain and a CD4 transmembrane domain; wherein ligand encoded by (a), ligand encoded by (b), and polypeptide encoded by (c) are fused directly to each other, or joined by at least one linker.
- CD19-TAC Trifunctional T cell-antigen coupler
- the nucleic acid sequence does not encode a co- stimulatory domain, an activation domain, or both a co-stimulatory domain and an activation domain.
- the ligand that selectively binds the CD 19 antigen is a single chain variable fragment (scFv).
- the ligand that selectively binds the CD 19 antigen comprises an amino acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 36.
- the ligand that selectively binds the CD19 antigen comprises an amino acid sequence of SEQ ID NO: 36.
- the huUCHTl (Y177T) ligand is a single chain antibody.
- the huUCHTl (Y177T) ligand comprises an amino acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 46. In some embodiments, the huUCHTl (Y177T) ligand comprises an amino acid sequence of SEQ ID NO: 46. In some embodiments, the third polynucleotide encodes a polypeptide comprising an amino acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 18.
- the third polynucleotide encodes a polypeptide comprising an amino acid sequence of SEQ ID NO: 18.
- the at least one linker is a G4S flexible linker, a large protein domain, a long helix structure, or a short helix structure.
- the at least one linker comprises an amino acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 12, SEQ ID NO: 32, SEQ ID NO: 30, or SEQ ID NO: 28.
- the at least one linker comprises an amino acid sequence of SEQ ID NO: 12, SEQ ID NO: 32, SEQ ID NO: 30, or SEQ ID NO: 28.
- the CD3 is expressed on a cell expressing the second polynucleotide.
- the CD19-TAC comprises a nucleic acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises an amino acid sequence having at least 80%, at least 90%, or at least 95% sequence identity with SEQ ID NO: 64.
- the CD19-TAC comprises a sequence of SEQ ID NO: 63 or SEQ ID NO: 64.
- vector constructs comprising: (a) a nucleic acid sequence disclosed herein; and (b) a promoter functional in a mammalian cell.
- compositions comprising the vector disclosed herein, and an excipient.
- nucleic acid sequences encoding a HER2 Trifunctional T cell-antigen coupler (HER2-TAC) comprising a nucleic acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 65, SEQ ID NO: 67, or SEQ ID NO:
- the HER2-TAC comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 66, SEQ ID NO: 68, or SEQ ID NO: 76.
- the nucleic acid sequence does not encode a co-stimulatory domain. In some embodiments, the nucleic acid sequence does not encode an activation domain.
- nucleic acid sequences encoding a BCMA Trifunctional T cell-antigen coupler comprising a nucleic acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 55, SEQ ID NO: 57, SEQ ID NO: 59, or SEQ ID NO: 61.
- the BCMA-TAC comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 60, or SEQ ID NO: 62.
- the nucleic acid sequence does not encode a co-stimulatory domain. In some embodiments, the nucleic acid sequence does not encode an activation domain.
- Fig. 1A is a schematic of natural T-cell activation.
- Fig. IB is a schematic of CAR based T-cell activation.
- Fig. 1C is a schematic of a trifunctional-T cell-antigen coupler (Tri-TAC) based T cell activation.
- Tri-TAC trifunctional-T cell-antigen coupler
- Fig. ID is a schematic of natural T-cell activation.
- Fig. IE is a schematic of CAR based T-cell activation.
- Fig. IF is a schematic of Tri-TAC based T cell activation.
- Fig. 2A is a schematic of a Tri-TAC configuration with the UCHT1 domain being centered between the trans-membrane domain (TM) and the antigen binding domain.
- Fig. 2B is a schematic of a Tri-TAC configuration in which the UCHT1 domain is N- terminal, followed by the antigen binding domain and the trans-membrane domain.
- Fig. 2C is a schematic of a Tri-TAC molecule with a generic antigen binding domain and a UCHTl domain.
- Fig. 3A is a schematic of a Tri-TAC molecule with a generic antigen binding domain.
- Fig. 3B is a schematic of a Tri-TAC with an anti-HER-2 DARPin antigen binding domain.
- Fig. 3C is a schematic of a Tri-TAC with an anti-CD 19 scFv antigen binding domain.
- Fig. 3D is a schematic of a Tri-TAC with an anti-BCMA scFv antigen binding domain.
- Fig. 3E is a schematic of a Tri-TAC molecule with the Anti-HER-2 DARPin antigen binding domain.
- Fig. 3F is a schematic of a Tri-TAC molecule with the Anti-BCMA scFv antigen binding domain.
- Fig. 4A-Fig. 4D exemplify T cells engineered with a Tri-TAC or a CD28-based CAR directed against HER-2 using a DARPin.
- Fig. 4A exemplifies the surface expression of the Tri- TAC and CAR compared to T cells that express no chimeric receptor.
- Fig. 4B exemplifies growth of three cell populations.
- Fig. 4C- Fig. 4D exemplify the percentage of engineered cells positive for various T cell activation markers following stimulation with antigen.
- Fig. 5 illustrates a model of the CD19-TAC protein structure.
- Fig. 6A-Fig. 6J illustrate receptor surface expression and activation of various anti- HER-2 DARPin Tri-TAC controls.
- T cells were engineered with a Tri-TAC variant that lacks the targeting element (-DARPin), a Tri-TAC variant that lacks UCHT1 (-UCHT1), or the full- length Tri-TAC.
- Fig. 6A, Fig. 6D, Fig. 6G illustrate T cell transduction and Her2 binding ability (left); Fig. 6B, Fig. 6E, Fig. 6H degranulation (middle) and Fig. 6C, Fig. 6F, Fig. 61 cytokine production (right).
- Fig. 6J illustrates that only full length anti-HER-2 DARPin Tri-TAC is able to elicit a cytotoxic response.
- Fig. 7A-Fig. 7C illustrate anti -tumor activity, toxicity, and cytokine production of T cells engineered with either the anti-HER-2 DARPin Tri-TAC or the anti-HER-2 DARPin CD28-based CAR.
- Mice bearing established OVCAR-3 tumors were treated with T cells engineered with the anti-HER-2 DARPin Tri-TAC or the anti-HER-2 DARPin CAR.
- Fig. 7A exemplifies the change in tumor growth relative to the day of T cell infusion (day 35).
- Fig. 7B exemplifies the change in weight, a measure of toxicity, in the same mice.
- Fig. 7C illustrates cytokine concentrations in serum of mice on day 7 post T-cell infusion.
- Fig. 8A-Fig. 8H illustrate Tri-TACs designed with various alternatives to the ETCHT1 scFv-CD3 recruitment domain.
- Fig. 8A provides a schematic representation of TAC receptor constructs utilizing the anti-HER-2 DARPin, paired with either the ETCHT1 or OKT3 anti-CD3 scFv.
- Fig. 8B illustrates HER-2 TAC surface expression of CD8+ NGFR+ (left) or CD4+ NGFR+ T cells (right).
- Fig. 8C, Fig. 8C1 illustrate cytokine production by HER-2-specific TAC-T cells stimulated with antigen-positive SK-OV-3 tumor cells.
- Fig. 8A-Fig. 8H illustrate Tri-TACs designed with various alternatives to the ETCHT1 scFv-CD3 recruitment domain.
- Fig. 8A provides a schematic representation of TAC receptor constructs utilizing the anti-HER-2 DARPin, paired with either the ETCHT1
- FIG. 8D illustrates killing of SK-OV-3 tumor cells by HER-2 TAC and vector control (vector only carrying tNGFR) T cells.
- Vector control T cells (circles) are compared against HER-2-specific TAC-T cells bearing ETCHT1 (square) or OKT3 (triangle).
- Fig. 8E provides a schematic representation of TAC receptor constructs utilizing the anti-CD 19 scFv, paired with either huUCHTl, F6A, or L2K anti-CD3 scFv.
- Fig. 8F illustrates CD19-TAC surface expression of CD8+ NGFR+ (left) or CD4+ NGFR+ T cells (right).
- FIG. 8G1 illustrate cytokine production by CD 19- specific TAC-T cells stimulated with antigen-positive Raji tumor cells. Cytokine producing cells are compared from TAC-T cells bearing huUCHTl (square), F6A (triangle), or L2K (diamond).
- Fig. 8H illustrates killing of NALM-6 tumor cells by CD19 TAC and vector control (vector only carrying tNGFR) T cells.
- Vector control T cells (circles) are compared against CD 19-specific TAC-T cells bearing huUCHTl (square), F6A (triangle), or L2K (diamond).
- Fig. 9A-Fig. 9H illustrates the effect of various anti-CD3 scFv on TCR surface expression.
- Fig. 9A, Fig. 9E illustrate TCR surface expression of T cells engineered with either control vector (tNGFR), UCHT1, or OKT3 TAC variants.
- Fig. 9B, Fig. 9F illustrate that T cells engineered with OKT3-TAC have significantly reduced TCR surface expression relative to UCHT1-TAC.
- Fig. 9C, Fig. 9G illustrate TCR surface expression of T cells engineered with control vector (tNGFR), huUCHTl, F6A or L2K TAC variants.
- Fig. 9D, Fig. 9H illustrates that T cells engineered with L2K TAC have significantly reduced TCR surface expression relative to huUCHTl -TAC.
- Fig. IOA-Fig. 10B illustrate connector domain variants.
- the domain the connecting antigen binding domain with the TCR recruitment domain is termed the connector domain.
- Fig. 10A provides schematics of TAC variants with different connector domains: (i) a flexible connector, (ii) a large domain connector (constructed from domains 3 and 4 derived from the extracellular CD4 domain), (iii) a long helical connector, and (iv) a short helical connector.
- Fig. 10B provides exemplary amino acid sequence of the domains represented in Fig. 10A. (SEQ ID NOS 69, 28, 30, and 32, respectively, in order of appearance)
- Fig. llA-Fig. 11E illustrate exemplary in vitro parameters of CD 19 TAC engineered with different connector variants.
- Fig. 11A illustrates TAC variant surface expression in both CD4 and CD8 cells.
- Fig. 11B illustrates surface expression of TAC comprising flexible connectors relative to TAC comprising helical or large domain connectors.
- Fig. 11C illustrates overall transduction of TAC comprising alternative connectors relative to the flexible connector.
- Fig. 11D, Fig. 11E illustrate relative cell reactivity to antigen positive Raji cells.
- Fig. 12A illustrates in vitro cytotoxicity of BCMA Tri-TAC variants engineered with different connectors.
- Fig.l2B illustrates in vivo tumor control of BCMA Tri-TAC variants engineered with the flexible connector compared to the short helical connector.
- Fig. 13A-Fig. 13C illustrate properties of CD8a Tri-TAC scFv anti HER-2, and CD8a Tri-TAC DARPin anti-HER-2.
- Fig. 13A, Fig. 13C illustrate surface expression.
- Fig. 13B illustrates cytokine production.
- Fig. 14A-Fig. 14D provide schematics of CD8 Tri-TAC variants.
- the anti HER-2- DARPin is used as an exemplary antigen-binding domain and the UCHT1 CD3 recruitment domain is used as an exemplary recruitment domain.
- Fig. 14A illustrates a Tri-TAC comprising a CD4 transmembrane and cytosolic domain (left), and comparable regions of a CD8o/CD8p heterodimer (right). Key regions for co-receptor functionality (arginine rich domain and CXCP motif) are highlighted.
- Fig. 14A illustrates a Tri-TAC comprising a CD4 transmembrane and cytosolic domain (left), and comparable regions of a CD8o/CD8p heterodimer (right). Key regions for co-receptor functionality (arginine rich domain and CXCP motif) are highlighted.
- FIG. 14B is a schematic of a CD8a Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a CD8a cytosolic domain.
- Fig. 14C is a schematic of a CD8a+R.p Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a chimeric CD8a cytosolic domain where the CD8a arginine rich region is replaced with the CD8P arginine rich region.
- 14D is a schematic of a CD8P+Lck Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a chimeric CD8P cytosolic domain, where the CD8a CXCP domain, which contains a Lck binding motif, was added to the C-terminus of the CD8P cytosolic domain.
- Fig. 15A-Fig. 15E illustrate in vitro characterization of CD8 Tri-TAC variants relative to the prototypic Tri-TAC containing CD4 regions.
- Fig. 15A-Fig. 15B illustrate surface expression of CD8-Tri TAC variants relative to the prototypic Tri-TAC.
- Fig. 15C illustrates in vitro cytotoxicity of CD8-Tri TAC variants co-cultured with LOX IMVI (HER-2 negative) or A549, SKOV3, SKBR3 or MBA MB 231 (HER-2 positive).
- Fig. 15D illustrates cell division of T cells engineered with either the CD8 Tri-TAC variants or the prototypic Tri-TAC.
- Fig. 15E illustrates TCR surface expression of engineered T cells comprising CD8 Tri-TAC variants or the prototypic Tri-TAC.
- Fig. 16 illustrates various Tri-TACs.
- Fig. 17 illustrates TAC-CD19 insert in a pCCL lentiviral vector.
- Fig. 17 illustrates the various domains of a TAC-CD19 (a CD8a leader, FMC63 scFv, Myc Tag, huUCHTl Y177T mutant and a truncated CD4 anchoring co-receptor domain).
- Fig. 18 illustrates the in vivo efficacy of TAC-CD19 generated from different donors.
- Fig. 19A-Fig. 19C illustrates an in vitro example of TAC-CD19 cytotoxicity against the tumor lines.
- Fig. 19C Raji Burkitt's lymphoma.
- Fig. 19D illustrates the schematic of 3 different in vivo tumor models in NRG mice.
- Fig. 19E - Fig. 19G illustrate in vivo efficacy of CD19-TAC in NALM-6 (acute lymphoblastic leukemia) Fig 19E, Jeko-l(Mantle Cell Lymphoma) Fig. 19F, and Raji (Burkitt's lymphoma) Fig. 19G.
- Fig. 20A illustrates the experimental set up of TAC-CD19 treated mice with NALM-6 tumor. Following successful treatment mice are then re-challenged with either NALM-6 (CD 19 positive) or KMS11 (CD 19 negative) tumor cells.
- Fig. 20B illustrates in vivo efficacy of mice treated with TAC-CD19.
- Fig. 21A illustrates the experimental design of evaluating dose regime and dosing impact on efficacy and cell expansion.
- Fig. 21B illustrates in vivo survival of NALM-6 bearing mice treated with either a single or split dose of TAC-CD19.
- Fig. 22A-Fig. 22B illustrate an experimental setup and data with regard to in vivo expansion of TAC-CD19 following a split dose administration.
- Fig. 22A illustrates the gating strategy used to identify T cells in mouse blood.
- Fig. 22B illustrates in vivo results of T cell expansion in blood.
- Fig. 23A - Fig. 23C illustrate long term in vivo studies of TAC-CD19 in mice.
- Fig. 23A illustrates an experimental protocol of NALM-6 bearing mice being treated with various controls and TAC-CD19 at two dose levels.
- Fig. 23B illustrates in vivo efficacy of control vs two dose levels of TAC-CD19 treatment groups.
- Fig. 23C illustrates long term survival of low dose TAC-CD19 treated mice.
- Fig. 24 illustrates clinical chemistry analysis results from mice treated with TAC-CD19 or non-transduced T cells.
- Fig. 25 illustrates human cytokine released in mice blood following treatment with TAC- CD19 or non-transduced T cells.
- Fig. 26A-Fig. 26C illustrates efficacy of BCMA-TAC in different configurations.
- Fig. 26A illustrates an experimental design.
- Fig. 26B illustrates various controls and test articles.
- Fig. 26C illustrates in vivo efficacy of various TAC constructs.
- Fig. 26A-Fig. 26C disclose "G 4 S” as SEQ ID NO: 73.
- Fig. 27 illustrates that TACs proliferate when encountering antigen on cells, but not when the antigen is presented on artificial beads; but CARs proliferate irrespective if antigens are presented on beads or cells.
- Fig. 28A-Fig. 28B illustrate TAC engineered T cells expand in vivo and provide long term protection, indicating cell persistence in a model of myeloma.
- Fig. 28A-Fig. 28B illustrate BCMA-TAC T cells reject multiple myeloma tumors in a KMS-l 1 xenograft model engineered with NanoLuc (KMS 1 l-NanoLuc) (BCMA pos ). Following tumor engraftment mice were treated with BCMA TAC-T cells (carrying Firefly Luciferase). TAC-T cells expand significantly following administration. This correlates with tumor regression. Treated mice were resistant to tumor rechallenge indicating long term persistence of TAC-T cells.
- Fig. 29 illustrates human cytokine released in mice blood following treatment with TAC- CD19 or non-transduced T cells.
- Fig. 30 illustrates exemplary histograms of TAC receptor surface expression in CD4 and CD8 engineered T cells.
- Cells were engineered with the muIgG HER2 TAC (EFla promoter), the huIgG HER2 TAC (MSCV promoter) and muIgG HER2 TAC (MSCV promoter).
- T cells were stained with a TAC specific reagent and measured using flow cytometry. All constructs show comparable levels of surface expression, with EFla driven expression being higher compared to the MSCV constructs.
- Fig. 31 illustrates the relative percentage of T cells expressing either TNFa, IFNy or IL-2 following co-cultured either with OVCAR3 (HER2 positive) or LOX IMVI (HER2 negative) cells.
- T cells were engineered with the muIgG HER2 TAC (EFla promoter), a TAC construct lacking the HER2 binding domain (Abinding TAC; EFla promoter), the huIgG HER2 TAC (MSCV promoter) or the muIgG HER2 TAC (MSCV promoter).
- LOX- IMVI HER2 neg
- OVCAR3(HER2 pos ) show similar ability to produce cytokines while control T cells engineered with Abinding TAC show no meaningful cytokine production.
- Fig 32 Illustrates in vivo efficacy of TAC engineered T cells in the OVCAR3 solid tumor model.
- T cells were engineered with the Abinding TAC (EFla promoter), the muIgG HER2 TAC (EFla promoter), the huIgG HER2 TAC (MSCV promoter) or the muIgG HER2 TAC (MSCV promoter).
- Mice had been inoculated subcutaneously with OVCAR3 (HER-2- positive) tumors. These were grown to about 100 mm 3 in size. Mice were then treated with a split dose of 6 million total HER2 TAC engineered, or Abinding TAC control T cells 48h apart via tail vain injection. Tumor progression was followed by biweekly measurements.
- Abinding TAC showed no tumor control or tumor regression. All HER2 TAC engineered T cells showed significantly reduced tumor progression, including tumor regression, relative to control mice.
- All HER2 TAC engineered T cells had similar anti-tumor activity.
- Cancer is a major health challenge, with over 150,000 cases of cancer expected to be diagnosed in Canada alone. While patients with early stage disease are sometimes treated effectively by conventional therapies (surgery, radiation, chemotherapy), few options are available to patients with advanced disease, and those options are typically palliative in nature.
- Active immunotherapy seeks to employ the patient’s immune system to clear tumors and offers an option to patients who have failed conventional therapies.
- this treatment involves infusing patients with large numbers of tumor-specific T cells.
- This approach has proven to be successful in early phase clinical trials for a number of diseases, including melanoma, myeloma, leukemia, lymphoma and synovial sarcoma.
- diseases including melanoma, myeloma, leukemia, lymphoma and synovial sarcoma.
- immunotherapy with T cells are curative in patients with advanced melanoma, confirming the utility of this approach.
- CLL chronic lymphocytic leukemia
- ALL acute lymphoblastic leukemia
- T cells isolated from a tumor-bearing patient are grown to large numbers ex vivo and are administered back into the patient to induce a robust anti-tumor immune response.
- Tumor specificity is achieved by either: (i) isolating naturally-occurring
- tumor-specific T cells from the patient; or (ii) engineering bulk T cells from the peripheral blood to express tumor-specific receptors.
- Naturally occurring tumor-specific T cells are rare and isolating such cells in therapeutic quantities from cancer patients is a laborious and costly procedure.
- T cell receptor TCR
- CAR chimeric antigen receptor
- the chimeric antigen receptors used for engineering T cells consist of: (i) a targeting domain, usually a single-chain fragment variable (scFv); (ii) a transmembrane domain; and (iii) a cytosolic domain that contains signaling elements from the T cell receptor and associated proteins.
- scFv single-chain fragment variable
- Such chimeric antigen receptors have also been referred to as“T-body” or“Chimeric Immune Receptor” (CIR), but currently, most researchers use the term“CAR”.
- CIR Choimeric Immune Receptor
- One advantage of the CAR approach is that it allows any patient’s immune cells to be targeted against any desirable target in a major histocompatibility complex (MHC) independent manner. This is appealing as MHC presentation is often defective in tumor cells.
- MHC major histocompatibility complex
- CARs are considered in modular terms and scientists have spent considerable time investigating the influence of different cytoplasmic signaling domains on CAR function.
- CARs generally share two main components: (i) the CD3 zeta cytoplasmic domain, which contains immunotyrosine activation motifs (ITAMs) critical for T cell activation; and (ii) components of costimulatory receptors that trigger important survival pathways such as the Akt pathway.
- the first-generation CARs employed a single signaling domain from either 003z or FceRTy.
- Second-generation CARs combined the signaling domain of CD3z with the cytoplasmic domain of costimulatory receptors from either the CD28 or TNFR family of receptors.
- CAR-engineered T cells that are currently being tested in the clinic employ second-generation CARs where 003z is coupled to the cytoplasmic domain of either CD28 or CD137. These second generation CARs have demonstrated anti -turn or activity in CD 19-positive tumors. Third- generation CARs combined multiple costimulatory domains, but there is concern that third- generation CARs may lose antigen-specificity.
- TCR T cell receptor
- BiTEs Bispecific T-cell Engagers
- These proteins employ bispecific antibody fragments to crosslink T- cell TCR receptors with target antigens. This leads to efficient T-cell activation, triggering cytotoxicity.
- bi-specific antibodies have been generated that accomplish this goal and some scientists have simply linked anti-CD3 antibodies to tumor-specific antibodies employing chemical linkage. While these bi-specific proteins have demonstrated some activity in vitro , GMP production, short biological half-lives, and limited bioavailability represent significant challenges to the successful use of these molecules in cancer treatment. Additionally, these molecules also fail to properly recapitulate natural TCR signaling because they do not engage the TCR co-receptors (CD8 and CD4).
- Tri-TAC Trifunctional T cell Antigen Coupler
- TAC Trifunctional T cell Antigen Coupler
- T cell refers to a type of lymphocyte that plays a central role in cell-mediated immunity.
- T cells also referred to as T lymphocytes, are distinguished from other lymphocytes, such as B cells and natural killer cells, by the presence of a T-cell receptor (TCR) on the cell surface.
- TCR T-cell receptor
- T cell antigen coupler or TAC is used interchangeably with“trifunctional T cell antigen coupler” or Tri-TAC and refers to an engineered nucleic acid construct or polypeptide, that when expressed on a T cell, helps to facilitate the targeting of the T cell to a particular antigen.
- the TAC comprises (a) a target-specific ligand, (b) a ligand that binds a protein associated with a T cell receptor (TCR) complex, and (c) a T cell receptor signaling domain.
- nucleic acid sequences of the present application may be deoxyribonucleic acid sequences (DNA) or ribonucleic acid sequences (RNA) and may include naturally occurring bases including adenine, guanine, cytosine, thymidine and uracil. The sequences may also contain modified bases.
- modified bases include aza and deaza adenine, guanine, cytosine, thymidine and uracil; and xanthine and hypoxanthine.
- the nucleic acids of the present disclosure may be isolated from biological organisms, formed by laboratory methods of genetic recombination or obtained by chemical synthesis or other known protocols for creating nucleic acids.
- isolated polynucleotide or“isolated nucleic acid sequence” as used herein refers to a nucleic acid substantially free of cellular material or culture medium when produced by recombinant DNA techniques, or chemical precursors, or other chemicals when chemically synthesized.
- An isolated nucleic acid is also substantially free of sequences which naturally flank the nucleic acid (i.e. sequences located at the 5' and 3' ends of the nucleic acid) from which the nucleic acid is derived.
- the term“nucleic acid” is intended to include DNA and RNA and is either double stranded or single stranded, and represents the sense or antisense strand. Further, the term“nucleic acid” includes the complementary nucleic acid sequences.
- recombinant nucleic acid or“engineered nucleic acid” as used herein refers to a nucleic acid or polynucleotide that is not found in a biological organism.
- recombinant nucleic acids may be formed by laboratory methods of genetic recombination (such as molecular cloning) to create sequences that would not otherwise be found in nature.
- Recombinant nucleic acids may also be created by chemical synthesis or other known protocols for creating nucleic acids.
- polypeptide or“protein” as used herein describes a chain of amino acids.
- a polypeptide or protein of this disclosure is a peptide, which usually describes a chain of amino acids.
- protein as used herein also describes a large molecule comprising one or more chains of amino acids and, in some embodiments, is a fragment or domain of a protein or a full length protein.
- protein either refers to a linear chain of amino acids or to a chain of amino acids that has been processed and folded into a functional protein.
- the protein structure is divided into four distinct levels: (1) primary structure - referring to the sequence of amino acids in the polypeptide chain, (2) secondary structure - referring to the regular local sub -structures on the polypeptide backbone chain, such as a-helix and b-sheets, (3) tertiary structure - referring to the three-dimensional structure if monomeric and multimeric protein molecules, and (4) quaternary structure - referring to the three-dimensional structure comprising the aggregation of two or more individual polypeptide chains that operate as a single functional unit.
- the proteins of the present disclosure are obtained by isolation and purification of the proteins from cells where they are produced naturally, by enzymatic (e.g., proteolytic) cleavage, and/or recombinantly by expression of nucleic acid encoding the proteins or fragments of this disclosure.
- the proteins and/or fragments of this disclosure in some embodiments, is obtained by chemical synthesis or other known protocols for producing proteins and fragments.
- isolated polypeptide refers to a polypeptide substantially free of cellular material or culture medium when produced by recombinant DNA techniques, or chemical precursors or other chemicals when chemically synthesized.
- antibody as used herein is intended to include monoclonal antibodies, polyclonal antibodies, single chain antibodies, chimeric antibodies, and antibody fusions.
- the antibody may be from recombinant sources and/or produced in transgenic animals.
- antibody fragment as used herein is intended to include without limitations Fab, Fab', F(ab')2, scFv, dsFv, ds-scFv, dimers, minibodies, diabodies, and multimers thereof, multispecific antibody fragments and Domain Antibodies.
- a vector refers to a polynucleotide that is used to deliver a nucleic acid to the inside of a cell.
- a vector is an expression vector comprising expression control sequences (for example, a promoter) operatively linked to a nucleic acid to be expressed in a cell.
- Expression control sequences for example, a promoter
- Vectors known in the art include, but are not limited to, plasmids, phages, cosmids and viruses.
- tumor antigen or“tumor associated antigen” as used herein refers to an antigenic substance produced in tumor cells that triggers an immune response in a host (e.g. which is presented by MHC complexes).
- a tumor antigen is on the surface of a tumor cell.
- T cell receptor refers to a complex of integral membrane proteins that participates in the activation of T cells in response to the binding of an antigen.
- the TCR is a disulfide-linked membrane-anchored heterodimer normally consisting of the highly variable alpha (a) and beta (b) chains expressed as part of a complex with the invariant CD3 (cluster of differentiation 3) chain molecules.
- T cells expressing this receptor are referred to as a:b (or ab) T cells, though a minority of T cells express an alternate receptor, formed by variable gamma (g) and delta (d) chains, referred as gd T cells.
- CD3 is a protein complex composed of four distinct chains. In mammals, the complex contains a CD3y chain, a CD3b chain, two CD3e chains and two O ⁇ 3z chains.
- transmembrane and cytosolic domain refers to a polypeptide that comprises a transmembrane domain and a cytosolic domain of a protein associated with the T cell receptor (TCR) complex.
- TCR T cell receptor
- transmembrane and cytosolic domain may include, but is not limited to, protein domains that (a) associate with the lipid raft and/or (b) bind Lck.
- A“TCR co-receptor” as used herein, refers to a molecule that assists the T cell receptor (TCR) in communicating with an antigen-presenting cell and may be considered part of the first signal that leads to the activation of the TCR.
- TCR co-receptors include, but are not limited to, CD4, LAG3, and CD8.
- A“TCR co-stimulator” as used herein, refers to a molecule that enhances the response of a T cell to an antigen and may be considered as the second signal that leads to the activation of the TCR.
- TCR co-stimulators include, but are not limited to, ICOS, CD27, CD28, 4-1BB (CD 137), 0X40 (CD134), CD30, CD40, lymphocyte fiction-associated antigen 1 (LFA- 1), CD2, CD7, LIGHT, NKG2C, B7-H3, and a ligand that specifically binds CD83.
- A“TCR co-inhibitor” or“checkpoint receptor” as used herein, refers to a molecule that inhibits the response of a T cell to an antigen.
- TCR co-inhibitors include, but are not limited to, PD-l, TIM3, LAG-3, TIGIT, BTLA, CD160, and CD37.
- the terms“recipient”,“individual”,“subject”,“host”, and“patient”, are used interchangeably herein and in some embodiments, refer to any mammalian subject for whom diagnosis, treatment, or therapy is desired, particularly humans.
- “Mammal” for purposes of treatment refers to any animal classified as a mammal, including humans, domestic and farm animals, and laboratory, zoo, sports, or pet animals, such as dogs, horses, cats, cows, sheep, goats, pigs, mice, rats, rabbits, guinea pigs, monkeys etc. In some embodiments, the mammal is human. None of these terms require the supervision of medical personnel.
- the terms“treatment,”“treating,” and the like refer to administering an agent, or carrying out a procedure, for the purposes of obtaining an effect.
- the effect may be prophylactic in terms of completely or partially preventing a disease or symptom thereof and/or may be therapeutic in terms of affecting a partial or complete cure for a disease and/or symptoms of the disease.
- Treatment may include treatment of a disease or disorder (e.g.
- cancer in a mammal, particularly in a human, and includes: (a) preventing the disease or a symptom of a disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it (e.g., including diseases that may be associated with or caused by a primary disease; (b) inhibiting the disease, i.e., arresting its development; and (c) relieving the disease, i.e., causing regression of the disease.
- Treating may refer to any indicia of success in the treatment or amelioration or prevention of a cancer, including any objective or subjective parameter such as abatement; remission; diminishing of symptoms; or making the disease condition more tolerable to the patient; slowing in the rate of degeneration or decline; or making the final point of degeneration less debilitating.
- the treatment or amelioration of symptoms is based on one or more objective or subjective parameters; including the results of an examination by a physician.
- treating includes the administration of the compounds or agents of the present invention to prevent, delay, alleviate, arrest or inhibit development of the symptoms or conditions associated with diseases (e.g. cancer).
- therapeutic effect refers to the reduction, elimination, or prevention of the disease, symptoms of the disease, or side effects of the disease in the subject.
- reference to a range of 90-100% includes 91%, 92%, 93%, 94%, 95%, 95%, 96%, 97%, etc., as well as 91.1%, 91.2%, 91.3%, 91.4%, 91.5%, etc., 92.1%, 92.2%, 92.3%, 92.4%, 92.5%, etc., and so forth.
- reference to a range of 1-5,000 fold includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, fold, etc., as well as 1.1, 1.2, 1.3, 1.4, 1.5, fold, etc., 2.1, 2.2, 2.3, 2.4, 2.5, fold, etc., and so forth.
- “About” a number refers to range including the number and ranging from 10% below that number to 10% above that number.“About” a range refers to 10% below the lower limit of the range, spanning to 10% above the upper limit of the range.
- “Percent (%) identity” refers to the extent to which two sequences (nucleotide or amino acid) have the same residue at the same positions in an alignment.
- “an amino acid sequence is X% identical to SEQ ID NO: Y” refers to % identity of the amino acid sequence to SEQ ID NO: Y and is elaborated as X% of residues in the amino acid sequence are identical to the residues of sequence disclosed in SEQ ID NO: Y.
- computer programs are employed for such calculations.
- Exemplary programs that compare and align pairs of sequences include ALIGN (Myers and Miller, 1988), FASTA (Pearson and Lipman, 1988; Pearson, 1990) and gapped BLAST (Altschul et al., 1997), BLASTP, BLASTN, or GCG (Devereux et al.,
- the term“selective binding” refers to the higher affinity with which a molecule (e.g. protein such as a target-binding ligand of TAC) binds its target molecule (e.g. target antigen such as HER-2, BCMA, or CD 19) over other molecules.
- a molecule e.g. protein such as a target-binding ligand of TAC
- target molecule e.g. target antigen such as HER-2, BCMA, or CD 19
- T cell antigen coupler Tri-TAC or TAC
- nucleic acids encoding a Trifunctional T cell-antigen coupler comprises: (a) a first polynucleotide encoding a target-specific ligand; (b) a second
- nucleic acids encoding a Tri-TAC do not encode a co-stimulatory domain. In some embodiments, the nucleic acids encoding a Tri-TAC do not encode a co-activation domain.
- the target-specific ligand also referred to as an antigen binding domain, refers to any substance or molecule that binds, directly or indirectly, to a target cell.
- the target specific ligand binds to an antigen on the target cell.
- a target cell is a cell associated with a disease state, including, but not limited to, cancer, hematologic malignancy, large B-cell lymphoma, diffuse large B-cell lymphoma, primary mediastinal B cell lymphoma, high grade B-cell lymphoma, or large B cell lymphoma arising from follicular lymphoma.
- a target cell is a tumor cell.
- a target-specific ligand binds to a tumor antigen or tumor associated antigen on a tumor cell.
- the target antigen is a tumor antigen.
- the tumor antigen when proteinaceous is a sequence of 8 or more amino acids up to the full protein.
- the tumor antigen is any number of amino acids in between 8 and the full length protein which comprises at least one antigenic fragment of the full length protein that is presented in a Major Histocompatibility Complex (MHC).
- MHC Major Histocompatibility Complex
- tumor antigens include, but are not limited to, CD 19, HER-2 (erbB-2), B-cell maturation antigen (BCMA),
- alphafetoprotein AFP
- carcinoembryonic antigen CEA
- CA-125 CA-125
- METC-l epithelial tumor antigen
- ETA epithelial tumor antigen
- MAGE melanoma-associated antigen
- PSA prostate-specific antigen
- glioma-associated antigen b-human chorionic gonadotropin, thyroglobulin, RAGE-l
- MN-CA IX human telomerase reverse transcriptase
- RET1, RET2 AS
- intestinal carboxyl esterase mut hsp70-2
- M-CSF prostase
- PAP NY-ESO-l
- LAGE-la p53
- prostein PSMA
- survivin and telomerase prostate-carcinoma tumor antigen-l (PCTA-l)
- ELF2M neutrophil elastase
- CD22 insulin growth factor (IGF)-I, IGF-II, IGF-I receptor
- the target-specific ligands include, but are not limited to, antibodies and fragments thereof, for example single chain antibodies such as single-chain antibodies (scFvs), single domain antibodies, peptides, peptidomimetics, proteins, glycoproteins, or proteoglycans that bind to the target cell and/or antigen.
- the target- specific ligands include, but are not limited to, designed ankyrin repeat proteins (DARPins), lectins, knottins, centryrins, anticalins, or naturally occurring ligands for the tumor antigen, such as growth factors, enzyme substrates, receptors or binding proteins.
- DARPins ankyrin repeat proteins
- target specific ligands include non-protein compounds that bind to target cells and/or antigens, including but not limited to carbohydrates, lipids, nucleic acids, or small molecules.
- a target-specific ligand is a designed ankyrin repeat (DARPin) targeted to a specific cell and/or antigen.
- DARPin ankyrin repeat
- a target-specific ligand is a single-chain variable fragment (ScFv) targeted to a specific cell and/or antigen.
- the tumor antigen is a HER-2 antigen.
- the HER-2 specific ligand comprises an antigen binding domain of an antibody selected from Trastuzumab, Pertuzumab, Lapatinib, Neratinib, Ado-trastuzmab Emtansine, Gancotamab, Margetuximab, Timigutuzumab, and Ertumaxomab.
- the target-specific ligand is a DARPin that selectively binds a HER-2 (erbB-2) antigen.
- the target-specific ligand is a DARPin that specifically binds a HER-2 (erbB-2) antigen.
- the DARPin targeted to HER-2 (erb-2) comprises SEQ ID NO: 7 or SEQ ID NO: 8
- the first polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 7. In some embodiments, the first
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 7. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 7. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 7. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 7. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 7. In some embodiments, the first polynucleotide comprises a nucleotide sequence of SEQ ID NO: 7.
- the target-specific ligand comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 8. In some embodiments, the target- specific ligand comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 8. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 8. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 8. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 8. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 8. In some embodiments, the target-specific ligand comprises an amino acid sequence of SEQ ID NO: 8.
- the tumor antigen is a BCMA antigen.
- the BCMA specific ligand comprises an antigen binding domain of an antibody selected from Belantamab mafodotin, and GSK2857916.
- the target-specific ligand is a scFv that selectively binds BCMA.
- the target-specific ligand is a scFv that specifically binds BCMA.
- the scFv that binds BCMA comprises SEQ ID NO: 33 or SEQ ID NO: 34.
- the first polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 33. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 33. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 33. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 33. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 33.
- the first polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 33. In some embodiments, the first polynucleotide comprises a nucleotide sequence of SEQ ID NO: 33.
- the target-specific ligand comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 34. In some embodiments, the target- specific ligand comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 34. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 34. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 34. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 34. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 34. In some embodiments, the target-specific ligand comprises an amino acid sequence of SEQ ID NO: 34.
- the tumor antigen is a CD 19 antigen.
- the target-specific ligand is a scFv that selectively binds CD 19.
- the target- specific ligand is a scFv that specifically binds CD 19.
- the scFv that binds CD19 comprises SEQ ID NO: 35 or SEQ ID NO: 36.
- the first polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 35. In some embodiments, the first
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 35. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 35. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 35. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 35. In some embodiments, the first polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 35.
- the first polynucleotide comprises a nucleotide sequence of SEQ ID NO: 35.
- the target-specific ligand comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 36. In some embodiments, the target- specific ligand comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 36. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 36. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 36.
- the target-specific ligand comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 36. In some embodiments, the target-specific ligand comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 36. In some embodiments, the target-specific ligand comprises an amino acid sequence of SEQ ID NO: 36.
- the TAC comprises a ligand that binds a protein associated with the TCR complex.
- the ligand that binds a protein associated with a TCR complex comprises a substance that binds, directly or indirectly, to a protein of the TCR.
- the ligand that binds a protein associated with a TCR complex comprises a substance that selectively binds to a protein of the TCR.
- the ligand that binds a protein associated with a TCR complex comprises a substance that specifically binds to a protein of the TCR.
- Proteins associated with the TCR include, but are not limited, to the TCR alpha (a) chain, TCR beta (b) chain, TCR gamma (g) chain, TCR delta (d) chain, CD3y chain, CD35 chain and CD3e chains.
- a ligand that binds a protein associated with the TCR complex is an antibody to the TCR alpha (a) chain, TCR beta (b) chain, TCR gamma (g) chain, TCR delta (d) chain, CD3y chain, CD35 chain and/or CD3e chain.
- the protein associated with a TCR complex is CD3.
- the protein associated with a TCR complex is CD3e.
- CD3 antibodies include, but are not limited to, for.
- the antibody that binds CD3 is a single chain antibody, for example a single-chain variable fragment (scFv).
- the ligand that binds a TCR is anti-CD3 antibody, or a fragment thereof, such as muromonab,
- otelixizumab otelixizumab
- teplizumab visilizumab
- CD3-12 MEM-57
- 4D10A6, CD3D or TR66.
- the CD3 is of a TCR complex on a cell expressing the second polynucleotide. In some embodiments, the binding of the CD3 induces activation of a cell expressing the second polynucleotide.
- the ligand that binds a TCR complex is UCHT1, or a variant thereof. In some embodiments, the ligand that binds a TCR complex is UCHT1 (SEQ ID NO:
- the UCHT1 ligand binds CD3. In some embodiments, the UCHT1 ligand selectively binds CD3. In some embodiments, the UCHT1 ligand specifically binds CD3. In some embodiments, the UCHT1 ligand binds CD3e.
- the UCHT1 ligand selectively binds CD3e. In some embodiments, the UCHT1 ligand specifically binds CD3e. In some embodiments, the UCHT1 ligand is encoded by SEQ ID NO 13. In some embodiments, the UCHT1 ligand comprises SEQ ID NO 14. In some embodiments, the UCHT1 ligand is mutated. In some embodiments, the UCHT1 ligand comprises a Y182T mutation (also referred to as UCHT1 (Y182T)) (SEQ ID NO: 71 and SEQ ID NO: 72). In some embodiments, the UCHT1 (Y182T) ligand binds CD3. In some embodiments, the UCHT1 ligand specifically binds CD3e. In some embodiments, the UCHT1 ligand is encoded by SEQ ID NO 13. In some embodiments, the UCHT1 ligand comprises SEQ ID NO 14. In some embodiments, the UCHT1 ligand is mut
- the UCHT1 (Y182T) ligand selectively binds CD3. In some embodiments, the UCHT1 (Y182T) ligand specifically binds CD3. In some embodiments, the UCHT1 (Y182T) ligand binds CD3e. In some embodiments, the UCHT1 (Y182T) ligand selectively binds CD3e. In some embodiments, the UCHT1 (Y182T) ligand specifically binds CD3e. In some
- the UCHT1 (Y182T) ligand is encoded by SEQ ID NO 71. In some embodiments, the UCHT1 (Y182T) ligand comprises SEQ ID NO 72. In some embodiments, the ligand that binds a TCR complex is a humanized UCHT1 (huUCHTl). In some embodiments, the ligand that binds a TCR complex is huUCHTl (SEQ ID NO 43, SEQ ID NO: 44 or homologs thereof). In some embodiments, the huUCHTl ligand binds CD3. In some embodiments, the huUCHTl ligand selectively binds CD3.
- the huUCHTl ligand specifically binds CD3. In some embodiments, the huUCHTl ligand binds CD3e. In some embodiments, the huUCHTl ligand selectively binds CD3e. In some embodiments, the huUCHTl ligand specifically binds CD3e. In some embodiments, the huUCHTl ligand is encoded by SEQ ID NO 43. In some embodiments, the huUCHTl ligand comprises SEQ ID NO 44. In some
- the huUCHTl has a Y177T mutation (also referred to as huUCHTl (Y177T)) (SEQ ID NO: 45 and SEQ ID NO: 46).
- the huUCHTl (Y177T) ligand binds CD3.
- the huUCHTl (Y177T) ligand selectively binds CD3.
- the huUCHTl (Y177T) ligand specifically binds CD3.
- the huUCHTl (Y177T) ligand binds CD3e. In some embodiments, the huUCHTl (Y177T) ligand selectively binds CD3e. In some embodiments, the huUCHTl (Y177T) ligand specifically binds CD3e. In some embodiments, the huUCHTl (Y177T) ligand is encoded by SEQ ID NO 45. In some embodiments, the huUCHTl ligand comprises SEQ ID NO 46.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 13. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 13. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 13. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 13. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 13.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 13. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 13.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 14. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 14. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 14. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 14. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 14.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 14. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 14.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 71. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 71. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 71. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 71.
- the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 71. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 71. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 71.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 72. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 72. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 72. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 72. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 72.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 72. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 72.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 43. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 43. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 43. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 43. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 43.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 43. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 43.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 44. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 44. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 44. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 44. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 44.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 44. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 44.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 45. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 45. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 45. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 45. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 45.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 45. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 45.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 46. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 46. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 46. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 46. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 46.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 46. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 46.
- the ligand that binds to a CD3 is OKT3.
- the OKT3 ligand binds CD3.
- the OKT3 ligand selectively binds CD3.
- the OKT3 ligand specifically binds CD3.
- the OKT3 ligand binds CD3e.
- the OKT3 ligand selectively binds CD3e.
- the OKT3 ligand specifically binds CD3e.
- the murine OKT3 ligand is encoded by SEQ ID NO 21.
- the OKT3 ligand comprises SEQ ID NO 22.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 21. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 21. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 21. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 21. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 21.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 21. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 21.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 22. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 22. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 22.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 22. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 22. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 22. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 22.
- the ligand that binds to a CD3 is F6A.
- the F6A ligand binds CD3.
- the F6A ligand selectively binds CD3.
- the F6A ligand specifically binds CD3.
- the F6A ligand binds CD3e.
- the F6A ligand selectively binds CD3e.
- the F6A ligand specifically binds CD3e.
- the murine F6A ligand is encoded by SEQ ID NO 23.
- the F6A ligand comprises SEQ ID NO 24.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 23. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 23. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 23. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 23. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 23.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 23. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 23.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 24. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 24. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 24. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 24. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 24.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 24. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 24.
- the ligand that binds to a CD3 is L2K. In some embodiments, the L2K ligand binds CD3. In some embodiments, the L2K ligand selectively binds CD3. In some embodiments, the L2K ligand specifically binds CD3. In some embodiments, the L2K ligand binds CD3e. In some embodiments, the L2K ligand selectively binds CD3e. In some
- the L2K ligand specifically binds CD3e.
- the murine L2K ligand is encoded by SEQ ID NO 25.
- the L2K ligand comprises SEQ ID NO 26.
- the second polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 25. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 25. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 25. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 25. In some embodiments, the second polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 25.
- the second polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 25. In some embodiments, the second polynucleotide comprises a nucleotide sequence of SEQ ID NO: 25.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 26. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 26. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 26. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 26. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 26.
- the ligand that binds a TCR complex comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 26. In some embodiments, the ligand that binds a TCR complex comprises an amino acid sequence of SEQ ID NO: 26.
- a T cell antigen coupler includes a T cell receptor signaling domain polypeptide.
- the TCR signaling domain polypeptide comprises a transmembrane domain.
- the TCR signaling domain polypeptide comprises a cytosolic domain.
- the TCR signaling domain polypeptide comprises a transmembrane domain and a cytosolic domain.
- the cytosolic domain and transmembrane domains are optionally joined by a linker.
- the T cell receptor signaling domain polypeptide comprises a TCR co-receptor domain.
- the T cell receptor signaling domain polypeptide does not comprise a TCR co-stimulator domain.
- the TCR signaling domain polypeptide comprises a transmembrane domain and/or a cytosolic domain of a TCR co receptor.
- the TCR co-receptor is CD4, CD8, LAG3, or a chimeric variation thereof.
- the TCR co-receptor is CD4.
- the TCR signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD4 co-receptor encoded by SEQ ID NO: 17.
- the TCR signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD4 co-receptor comprising SEQ ID NO: 18.
- the third polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 17. In some embodiments, the third
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 17. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 17. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 17. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 17. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 17. In some embodiments, the third polynucleotide comprises a nucleotide sequence of SEQ ID NO: 17.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 18. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 18. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 18. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 18. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 18.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 18. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence of SEQ ID NO: 18.
- the TCR co-receptor is CD8. In some embodiments, the TCR co- receptor is CD8a. In some embodiments, the TCR signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8a co-receptor encoded by SEQ ID NO: 37. In some embodiments, the TCR signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8a co-receptor comprising SEQ ID NO: 38.
- the third polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 37. In some embodiments, the third
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 37. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 37. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 37. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 37. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 37. In some embodiments, the third polynucleotide comprises a nucleotide sequence of SEQ ID NO: 37.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 38. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 38. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 38. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 38. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 38.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 38. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence of SEQ ID NO: 38.
- the TCR signaling domain polypeptide comprises a chimera of sequences or domains from co-receptors. In some embodiments, the TCR signaling domain polypeptide comprises a chimera of CD8a and CD8P, wherein the CD8a arginine rich region is replaced with the CD8P arginine rich region. In some embodiments, the T cell receptor signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8a+R(P) co receptor chimera encoded by SEQ ID NO: 39.
- the T cell receptor signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8a+R(P) co-receptor chimera provided by SEQ ID NO: 40.
- the TCR signaling domain polypeptide comprises a chimera of CD8a and CD8P, the CD8a CXCP domain, which contains an Lck binding motif, is appended to the C-terminus of the CD8P cytosolic domain.
- the T cell receptor signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8p+Lck co-receptor chimera encoded by SEQ ID NO: 41.
- the T cell receptor signaling domain polypeptide comprises the transmembrane and cytosolic domains of the CD8p+Lck co-receptor chimera provided by SEQ ID NO: 42.
- the third polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 39. In some embodiments, the third
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 39. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 39. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 39. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 39. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 39. In some embodiments, the third polynucleotide comprises a nucleotide sequence of SEQ ID NO: 39.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 40. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 40. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 40. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 40. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 40.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 40. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence of SEQ ID NO: 40.
- the third polynucleotide comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 41. In some embodiments, the third
- polynucleotide comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 41. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 41. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 41. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 41. In some embodiments, the third polynucleotide comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 41. In some embodiments, the third polynucleotide comprises a nucleotide sequence of SEQ ID NO: 41.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 42. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 42. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 42. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 42. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 42.
- the transmembrane domain and cytosolic domain comprise an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 42. In some embodiments, the transmembrane domain and cytosolic domain comprise an amino acid sequence of SEQ ID NO: 42.
- the T cell receptor signaling domain polypeptide comprises a TCR co-stimulator domain.
- the TCR co-stimulator is ICOS.
- the TCR co-stimulator is CD27.
- the TCR co-stimulator is CD28.
- the TCR co-stimulator is 4-1BB (CD137).
- the TCR co-stimulator is 0X40 (CD 134).
- the TCR co-stimulator is CD30.
- the TCR co-stimulator is CD40.
- the TCR co-stimulator is lymphocyte fiction-associated antigen 1 (LFA-l).
- the TCR co-stimulator is CD2. In some embodiments, the TCR co-stimulator is CD7. In some embodiments, the TCR co-stimulator is LIGHT. In some embodiments, the TCR co-stimulator is NKG2C. In some embodiments, the TCR co-stimulator is B7-H3. In some embodiments, the TCR co-stimulator is a ligand that specifically binds CD83.
- the TCR signaling domain polypeptide comprises a
- the TCR co-inhibitor is PD-l. In some embodiments, the TCR co-inhibitor is TIM3. In some embodiments, the TCR co-inhibitor is LAG-3. In some embodiments, the TCR co-inhibitor is TIGIT. In some embodiments, the TCR co-inhibitor is BTLA. In some embodiments, the TCR co-inhibitor is CD160. In some embodiments, the TCR co-inhibitor is CD37.
- the TCR signaling domain polypeptide includes both a cytosolic domain and a transmembrane domain of a TCR co-receptor or co-stimulator protein.
- the cytosolic domain and transmembrane domain are from the same co-receptor or co-stimulator or from different co-receptors or co-stimulators.
- the TAC further comprises other polypeptides that directly or indirectly act to target or activate the T cell.
- a nucleic acid disclosed herein is in an order of (1) a first polynucleotide encoding a target-specific ligand; (2) a second polynucleotide encoding a ligand that binds a TCR complex; (3) a third polynucleotide encoding a transmembrane domain and a cytosolic domain.
- a nucleic acid disclosed herein is in an order of (1) a first polynucleotide encoding a target-specific ligand; (2) a second polynucleotide encoding a ligand that binds a TCR complex; (3) a third polynucleotide encoding a transmembrane domain and a cytosolic domain, wherein the order is 5’ end to 3’ end.
- a nucleic acid disclosed herein is in an order of (1) a first polynucleotide encoding a target-specific ligand; (2) a second polynucleotide encoding a ligand that binds a TCR complex; (3) a third
- a nucleic acid described herein is in an order of (1) a first polynucleotide encoding a ligand that binds a TCR complex; (2) a second polynucleotide encoding a target-specific ligand; (3) a third polynucleotide encoding a transmembrane domain and a cytosolic domain.
- a nucleic acid described herein is in an order of (1) a first polynucleotide encoding a ligand that binds a TCR complex; (2) a second polynucleotide encoding a target-specific ligand; (3) a third polynucleotide encoding a transmembrane domain and a cytosolic domain, wherein the order is 5’ end to 3’ end.
- a nucleic acid described herein is in an order of (1) a first polynucleotide encoding a ligand that binds a TCR complex; (2) a second polynucleotide encoding a target- specific ligand; (3) a third polynucleotide encoding a transmembrane domain and a cytosolic domain, wherein the order is 3’ end to 5’ end.
- the first nucleic acid encodes a first polypeptide
- the second nucleic acid encodes a second polypeptide
- the third nucleic acid encodes a third
- the first polypeptide, the second polypeptide, and the third polypeptide are directly fused.
- the target-specific ligand and the T cell receptor signaling domain polypeptide are both fused to the ligand that binds the TCR complex.
- the first polypeptide, the second polypeptide, and the third polypeptide are joined by at least one linker.
- the first polypeptide and the second polypeptide are directly fused, and joined to the third polypeptide by a linker.
- the second polypeptide and the third polypeptide are directly fused, and joined to the first polypeptide by a linker.
- the linker is a peptide linker. In some embodiments, the peptide linker comprises 1 to 40 amino acids. In some embodiments, the peptide linker comprises 1 to 30 amino acids. In some embodiments, the peptide linker comprises 1 to 15 amino acids. In some embodiments, the peptide linker comprises 1 to 10 amino acids. In some embodiments, the peptide linker comprises 1 to 6 amino acids. In some embodiments, the peptide linker comprises 30 to 40 amino acids. In some embodiments, the peptide linker comprises 32 to 36 amino acids. In some embodiments, the peptide linker comprises 5 to 30 amino acids. In some embodiments, the peptide linker comprises 5 amino acids.
- the peptide linker comprises 10 amino acids. In some embodiments, the peptide linker comprises 15 amino acids. In some embodiments, the peptide linker comprises 20 amino acids. In some embodiments, the peptide linker comprises 25 amino acids. In some embodiments, the peptide linker comprises 30 amino acids.
- the peptide linker comprises a G 4 S 3 linker (SEQ ID NO: 74). In some embodiments, the peptide linker comprises SEQ ID NOs: 11, 12, 15, 16, 19, 20, or variants or fragments thereof.
- the peptide linker that joins the target-specific ligand to the ligand that binds a TCR complex (e.g. UCHT1) is known as the connector to distinguish this protein domain from other linkers in the Tri-TAC.
- the connector is of any size.
- the connector between ligand that binds a TCR complex and a target-specific ligand is a short helix comprising SEQ ID NO ID: 28.
- the connector between ligand that binds a TCR complex and a target-specific ligand is a short helix encoded by SEQ ID NO ID:
- the connector between ligand that binds a TCR complex and a target- specific ligand is a long helix comprising SEQ ID NO ID: 30. In some embodiments, the connector between ligand that binds a TCR complex and a target-specific ligand is a long helix encoded by SEQ ID NO ID: 29. In some embodiments, the connector between ligand that binds a TCR complex and a target-specific ligand is a large domain comprising SEQ ID NO ID: 32. In some embodiments, the connector between ligand that binds a TCR complex and a target- specific ligand is a large domain encoded by SEQ ID NO ID: 31.
- a nucleic acid disclosed herein comprises a leader sequence.
- the leader sequence comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 5, 47, or 49.
- the leader sequence comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 5, 47, or 49.
- the leader sequence comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 5, 47, or 49.
- the leader sequence comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 5, 47, or 49.
- the leader sequence comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 5, 47, or 49. In some embodiments, the leader sequence comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 5, 47, or 49. In some embodiments, the leader sequence comprises a nucleotide sequence of SEQ ID NO: 5, 47, or 49.
- a nucleic acid disclosed herein comprises a leader sequence.
- the leader sequence comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 6, 48, or 50. In some embodiments, the leader sequence comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 6, 48, or 50. In some embodiments, the leader sequence comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 6, 48, or 50. In some embodiments, the leader sequence comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 6, 48, or 50. In some embodiments, the leader sequence comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 6, 48, or 50.
- the leader sequence comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 6, 48, or 50. In some embodiments, the leader sequence comprises an amino acid sequence of SEQ ID NO: 6, 48, or 50.
- the Tri-TAC is contemplated to be present in various configurations and combinations of (a) target-specific ligand, (b) a ligand that binds a TCR complex, and (c) a TCR signaling domain, as disclosed herein.
- the Tri-TAC comprises (a) a target-specific ligand, (b) a single- chain antibody (scFv) that binds CD3e, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a target-specific ligand, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a target-specific ligand, (b) F6A, and (c) a
- the Tri- TAC comprises (a) a target-specific ligand, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a DARPin, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a DARPin, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a DARPin,
- the Tri-TAC comprises (a) a DARPin, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a DARPin, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a DARPin, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri- TAC comprises (a) a DARPin, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a scFv, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a scFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a scFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a target-specific ligand, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri- TAC comprises (a) a HER-2-specific DARPin, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) UCHT1 (Y182T), and (c) a
- the Tri- TAC comprises (a) a BCMA-specific ScFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri- TAC comprises (a) a BCMA-specific ScFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a CDl9-specific ScFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a CD 19-specific ScFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor. In some embodiments, the Tri-TAC comprises (a) a CD 19-specific ScFv, (b) F6A, and (c) a
- the Tri- TAC comprises (a) a CD 19-specific ScFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) a single chain antibody (scFv) that binds CD3e, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri- TAC comprises (a) a target-specific ligand, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri-TAC comprises (a) a target-specific ligand, (b) F6A, and (c) a
- the Tri- TAC comprises (a) a target-specific ligand, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a DARPin, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD4 co-receptor.
- the Tri- TAC comprises (a) a DARPin, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a DARPin,
- the Tri-TAC comprises (a) a DARPin, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri- TAC comprises (a) a DARPin, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a DARPin, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri- TAC comprises (a) a DARPin, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri- TAC comprises (a) a scFv, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) huUCHTl (Y177T), and (c) a
- the Tri- TAC comprises (a) a scFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a scFv, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri- TAC comprises (a) a scFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) UCHT1 (Y182T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a target-specific ligand, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a HER-2-specific DARPin, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri- TAC comprises (a) a HER-2-specific DARPin, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) UCHT1 (Y182T), and (c) a
- the Tri- TAC comprises (a) a BCMA-specific ScFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri-TAC comprises (a) a BCMA-specific ScFv, (b) F6A, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri- TAC comprises (a) a BCMA-specific ScFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) UCHT1, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) UCHT1 (Y182T), and (c) a
- the Tri- TAC comprises (a) a CD 19-specific ScFv, (b) huUCHTl, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a CDl9-specific ScFv, (b) huUCHTl (Y177T), and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC comprises (a) a CD 19-specific ScFv, (b) OKT3, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor. In some embodiments, the Tri-TAC comprises (a) a CD 19-specific ScFv, (b) F6A, and (c) a
- the Tri- TAC comprises (a) a CD 19-specific ScFv, (b) L2K, and (c) a transmembrane and cytosolic domain of the CD8 co-receptor.
- the Tri-TAC draws CD3 and TCR into lipid raft regions of the membrane, and brings Lck into the proximity of the TCR, similar to natural MHC binding.
- the TAC disclosed herein is the anti-HER-2 DARPin Tri-TAC (also referred to as configuration 1; SEQ ID NO: 1 and 2) includes, in order:
- the TAC disclosed herein is a HER2-TAC.
- the HER2-TAC comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 65.
- the HER2-TAC comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 65.
- the HER2- TAC comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 65.
- the HER2-TAC comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 65.
- the HER2-TAC comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 65. In some embodiments, the HER2-TAC comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 65. In some embodiments, the HER2-TAC comprises a nucleotide sequence of SEQ ID NO: 65.
- the HER2-TAC comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 66. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 66. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 66. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 66. In some embodiments, the HER2-TAC an amino acid sequence having at least 90% sequence identity with SEQ ID NO:
- the HER2-TAC comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 66. In some embodiments, the HER2-TAC comprises an amino acid sequence of SEQ ID NO: 66.
- the TAC disclosed herein is a HER2-TAC.
- the HER2-TAC comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 67.
- the HER2-TAC comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 67.
- the HER2- TAC comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO:
- the HER2-TAC comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 67. In some embodiments, the HER2-TAC comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 67. In some embodiments, the HER2-TAC comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 67. In some embodiments, the HER2-TAC comprises a nucleotide sequence of SEQ ID NO: 67.
- the HER2-TAC comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 68. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 68. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 68. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 68. In some embodiments, the HER2-TAC an amino acid sequence having at least 90% sequence identity with SEQ ID NO:
- the HER2-TAC comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 68. In some embodiments, the HER2-TAC comprises an amino acid sequence of SEQ ID NO: 68.
- the TAC disclosed herein is a HER2-TAC.
- the HER2-TAC comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 75.
- the HER2-TAC comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 75.
- the HER2- TAC comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO:
- the HER2-TAC comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 75. In some embodiments, the HER2-TAC comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 75. In some embodiments, the HER2-TAC comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 75. In some embodiments, the HER2-TAC comprises a nucleotide sequence of SEQ ID NO: 75.
- the HER2-TAC comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 76. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 76. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 76. In some embodiments, the HER2-TAC comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 76. In some embodiments, the HER2-TAC an amino acid sequence having at least 90% sequence identity with SEQ ID NO:
- the HER2-TAC comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 76. In some embodiments, the HER2-TAC comprises an amino acid sequence of SEQ ID NO: 76.
- the TAC disclosed herein is a BCMA-TAC. In some embodiments, the TAC disclosed herein is a BCMA-TAC. In some
- the BCMA-TAC comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 55, 57, 59 or 61. In some embodiments, the BCMA-TAC comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 55, 57, 59 or 61. In some embodiments, the BCMA-TAC comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 55, 57, 59 or 61. In some embodiments, the BCMA-TAC comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 55,
- the BCMA-TAC comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 55, 57, 59 or 61. In some embodiments, the BCMA-TAC comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 55, 57, 59 or 61. In some embodiments, the BCMA-TAC comprises a nucleotide sequence of SEQ ID NO: 55, 57, 59 or 61. [0175] In some embodiments, the BCMA-TAC comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 56, 58, 60 or 62.
- the BCMA-TAC comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 56, 58, 60 or 62. In some embodiments, the BCMA-TAC comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 56, 58, 60 or 62. In some embodiments, the BCMA-TAC comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 56, 58, 60 or 62. In some embodiments, the BCMA-TAC comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 56, 58, 60 or 62.
- the BCMA-TAC comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 56, 58, 60 or 62. In some embodiments, the BCMA- TAC comprises an amino acid sequence of SEQ ID NO: 56, 58, 60 or 62.
- the TAC disclosed herein is a CD19-TAC.
- the CD19-TAC comprises a nucleotide sequence having at least 70% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises a nucleotide sequence having at least 75% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises a nucleotide sequence having at least 85% sequence identity with SEQ ID NO: 63.
- the CD19-TAC comprises a nucleotide sequence having at least 90% sequence identity with SEQ ID NO: 63. In some embodiments, the CD19-TAC comprises a nucleotide sequence having at least 95% sequence identity with SEQ ID NO: 63. In some embodiments, the CD19-TAC comprises a nucleotide sequence of SEQ ID NO: 63.
- the CD19-TAC comprises an amino acid sequence having at least 70% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence having at least 75% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 64. In some embodiments, the CD19-TAC comprises an amino acid sequence of SEQ ID NO: 64.
- polypeptides and Vector Constructs [0178] Disclosed herein, in certain embodiments, are polypeptides encoded by the nucleic acid sequence as disclosed herein. Also disclosed herein, are vectors comprising the nucleic acid sequence as disclosed herein. In some embodiments, the vectors further comprise a promoter. In some embodiments, the promoter is functional in a mammalian cell. Promoters, regions of DNA that initiate transcription of a particular nucleic acid sequence, are well known in the art. A “promoter functional in a mammalian cell” refers to a promoter that drives expression of the associated nucleic acid sequence in a mammalian cell. A promoter that drives expression of a nucleic acid sequence is referred to as being“operably connected” to the nucleic acid sequence.
- a variety of delivery vectors and expression vehicles are employed to introduce nucleic acids described herein into a cell.
- polynucleotides comprised in a vector to provide a vector construct, also herein referred to as a vector.
- a vector comprising:
- b a second polynucleotide encoding a ligand that binds a protein associated with a TCR complex
- c. a third polynucleotide encoding a T cell receptor signaling domain polypeptide; and d. a promoter that is functional in a mammalian cell.
- the target of the target-specific ligand binds to HER-2, BCMA, or CD 19.
- the target-specific ligand is a DARPin that selectively binds a HER-2 (erbB-2) antigen.
- the target-specific ligand is a DARPin that specifically binds a HER-2 (erbB-2) antigen.
- the DARPin targeted to HER-2 (erb-2) comprises SEQ ID NO: 7 or SEQ ID NO: 8.
- the target- specific ligand is a scFv that selectively binds BCMA.
- the target-specific ligand is a scFv that specifically binds BCMA.
- the scFv that binds BCMA comprises SEQ ID NO: 33 or SEQ ID NO: 34.
- the target-specific ligand is a scFv that selectively binds CD 19.
- the target-specific ligand is a scFv that specifically binds CD 19.
- the scFv that binds CD 19 comprises SEQ ID NO: 35 or SEQ ID NO: 36.
- the ligand that binds a protein associated with a TCR complex is UCHT1, humanized UCHT1 (huUCHTl), OKT3, F6A, or L2K.
- the ligand that binds a protein associated with a TCR complex is UCHT1, or a variant thereof.
- the ligand that binds a protein associated with a TCR complex is UCHT1 and is encoded by SEQ ID NO: 13.
- the ligand that binds a protein associated with a TCR complex is UCHT1 and comprises SEQ ID NO: 14
- the UCHT1 ligand that binds a protein associated with a TCR complex has a Y182T mutation (UCHT1 (Y182T)) and is encoded by SEQ ID NO: 71.
- the ligand that binds a protein associated with a TCR complex is UCHT1 (Y182T) and comprises SEQ ID NO: 72.
- the ligand that binds a protein associated with a TCR complex is humanized UCHT1 (huUCHTl), or a variant thereof.
- the ligand that binds a protein associated with a TCR complex is humanized UCHT1 (huUCHTl) and is encoded by SEQ ID NO: 43.
- the ligand that binds a protein associated with a TCR complex is huUCHTl and comprises SEQ ID NO: 44.
- the huUCHTl ligand that binds a protein associated with a TCR complex has a Y177T mutation (huUCHTl (Y177T)) and is encoded by SEQ ID NO: 45.
- the ligand that binds a protein associated with a TCR complex is huUCHTl (Y177T) and comprises SEQ ID NO: 46.
- the ligand that binds a protein associated with a TCR complex is OKT3, or a variant thereof.
- the ligand that binds a protein associated with a TCR complex is OKT3 and is encoded by SEQ ID NO: 21.
- the ligand that binds a protein associated with a TCR complex is OKT3 and comprises SEQ ID NO: 22
- the ligand that binds a protein associated with a TCR complex is F6A, or a variant thereof. In some embodiments, the ligand that binds a protein associated with a TCR complex is F6A and is encoded by SEQ ID NO: 23. In some embodiments, the ligand that binds a protein associated with a TCR complex is F6A and comprises SEQ ID NO: 24.
- the ligand that binds a protein associated with a TCR complex is L2K, or a variant thereof. In some embodiments, the ligand that binds a protein associated with a TCR complex is L2K and is encoded by SEQ ID NO: 25. In some embodiments, the ligand that binds a protein associated with a TCR complex is L2K and comprises SEQ ID NO: 26.
- the protein associated with a TCR complex is CD3. In some embodiments, the protein associated with a TCR complex is CD3e.
- the TCR signaling domain polypeptide comprises a
- the TCR co-receptor is CD4, CD8, LAG3, or a chimeric variation thereof.
- the first polynucleotide and third polynucleotide are fused to the second polynucleotide and the coding sequence is operably connected to the promoter.
- the second polynucleotide and third polynucleotide are fused to the first polynucleotide and the coding sequence is operably connected to the promoter.
- the vector is designed for expression in mammalian cells such as T cells.
- the vector is a viral vector.
- the viral vector is a retroviral vector.
- vectors that are useful comprise vectors derived from lentiviruses, Murine Stem Cell Viruses (MSCV), pox viruses, oncoretroviruses, adenoviruses, and adeno- associated viruses.
- Other delivery vectors that are useful comprise vectors derived from herpes simplex viruses, transposons, vaccinia viruses, human papilloma virus, Simian
- immunodeficiency viruses HTLV, human foamy virus and variants thereof.
- Further vectors that are useful comprise vectors derived from spumaviruses, mammalian type B retroviruses, mammalian type C retroviruses, avian type C retroviruses, mammalian type D retroviruses and HTLV/BLV type retroviruses.
- a lentiviral vector useful in the disclosed compositions and methods is the pCCL4 vector.
- the nucleic acid is a recombinant, or engineered, nucleic acid.
- the first, second and/or third polynucleotides are recombinant, or engineered, polynucleotides.
- the polynucleotides described herein are be modified or mutated to optimize the function of the encoded polypeptide and/or the function, activity and/or expression of the T cell antigen coupler.
- the nucleic acid encodes a polypeptide.
- modifications are made to the polynucleotide sequences including vector sequences and polypeptides sequences disclosed herein. Modifications include substitution, insertion or deletion of nucleotides or amino acids or altering the relative positions or order of nucleotides or amino acids.
- T cells comprising the nucleic acid sequences disclosed herein, or the vectors disclosed herein.
- human T cells engineered to express a Tri-TAC disclosed herein.
- the T cell expresses a Tri-TAC disclosed herein.
- the T cell is an isolated T cell.
- the human T cells engineered to express a Tri-TAC demonstrate functionality equivalent to a conventional CAR in vitro. In some embodiments, T cells engineered with the Tri-TAC demonstrate functionality superior to a conventional CAR in vitro. Disclosed herein, in some embodiments, are human T cells engineered with a Tri-TAC that demonstrate enhanced safety compared to traditional CARs. In some embodiments, human T cells engineered to express a Tri-TAC demonstrate enhanced safety compared to traditional CARs.
- T cells are obtained from a number of sources, including, but not limited to blood (for example, peripheral blood mononuclear cells), bone marrow, thymus tissue, lymph node tissue, cord blood, thymus tissue, tissue from an infection site, spleen tissue, or tumors.
- the T cells are autologous T cells.
- the T cells are obtained from a cell line of T cells.
- the T cells are obtained from donors (allogeneic T cells).
- the T cells are obtained by differentiation of embryonic or adult stem cells or from induced pluripotent stem cells.
- the T cells regardless of the source of T cells, the T cells have been modified so that they lack expression of an endogenous TCR and/or permanently or transiently lack expression of MHC/HLA molecules (universal donor T cells).
- the T cells are autologous with respect to the subject.
- the cells are allogeneic, syngeneic, or xenogeneic with respect to the subject.
- the T cells are optionally enriched in vitro.
- a population of cells is enriched by positive or negative selection.
- the T cells are optionally frozen or cryopreserved and then thawed at a later date.
- T cells are activated and/or expanded before or after introducing the Tri-TAC to the T cells.
- the T cells are expanded by contact with a surface having attached thereto an agent that stimulates a CD3/TCR complex associated signal and a ligand that stimulates a co-stimulator molecule on the surface of the T cells.
- the T cells are expanded by contact with one or more soluble agents that stimulate CD3/TCR complex signaling and co-stimulator molecule signaling.
- the T cells are transduced or transfected with nucleic acid sequences.
- the transduced or transfected T cells express proteins coded for by the transfected or transduced nucleic acid sequences.
- a nucleic acid may be introduced into a cell by physical, chemical, or biological means. Physical means include, but are not limited to, microinjection, electroporation, particle bombardment, lipofection and calcium phosphate precipitation.
- Bio means include the use of DNA and RNA vectors.
- Viral vectors are used to introduce and express a nucleic acid into a T cell.
- Viral vectors include vectors derived from lentivirus, Murine Stem Cell Viruses (MSCV), pox viruses, herpes simplex virus I, adenovirus and adeno-associated viruses.
- the vector optionally includes a promoter that drives expression of the transduced nucleic acid molecule in a T cell (e.g., a CMV promoter, eFla promoter, or MSCV promoter).
- Any suitable assay is used to confirm the presence and/or expression of the transduced nucleic acid sequence and/or the polypeptide encoded by the nucleic acid in the T cell. Assays include, but are not limited to, Southern and Northern blotting, RT-PCR and PCR, ELISA, Western blotting, and flow cytometry.
- a T cell expressing a TAC has increased T cell activation in the presence of an antigen compared to a T cell not expressing a TAC and/or as compared to a T cell expressing a traditional CAR.
- Increased T cell activation is ascertained by numerous methods, including but not limited to, increased tumor cell line killing, increased cytokine production, increased cytolysis, increased degranulation and/or increased expression of activation markers such as CDl07a, IFNy, IL2 or TNFa.
- increases are measured in an individual cell or in a population of cells.
- compositions comprising an engineered T cell disclosed herein (transduced with and/or expressing a TAC), and a pharmaceutically acceptable carrier.
- Pharmaceutically acceptable carriers include, but are not limited to, buffers such as neutral buffered saline, phosphate buffered saline and the like;
- the engineered T cells are formulated for intravenous administration.
- compositions are administered in a manner appropriate to the disease to be treated (or prevented).
- the quantity and frequency of administration is determined by such factors as the condition of the patient, and the type and severity of the patient’s disease, although appropriate dosages are determined by clinical trials.
- an immunologically effective amount “an anti-tumor effective amount,”“a tumor-inhibiting effective amount,” or “therapeutic amount” is indicated
- the precise amount of the compositions of the present invention to be administered is determined by a physician with consideration of individual differences in age, weight, tumor size, extent of infection or metastasis, and condition of the patient (subject).
- the modified T cells and/or pharmaceutical compositions described herein are administered at a dosage of 10 1 to 10 15 cells per kg body weight, 10 4 to 10 9 cells per kg body weight, optionally 10 5 to 10 8 cells per kg body weight, 10 6 to 10 7 cells per kg body weight or 10 5 to 10 6 cells per kg body weight, including all integer values within those ranges.
- the modified T cells and/or pharmaceutical compositions described herein are administered at a dosage of greater than 10 1 cells per kg body weight.
- the modified T cells and/or pharmaceutical compositions described herein are administered at a dosage of less than 10 15 cells per kg body weight.
- the modified T cells and/or pharmaceutical compositions described herein are administered at a dosage of 0.5 xlO 6 cells, 2 xlO 6 cells, 4 xlO 6 cells, 5 xlO 6 cells, 1.2 xlO 7 cells, 2 xlO 7 cells, 5 xlO 7 cells, 2 xlO 8 cells, 5 xlO 8 cells, 2 xlO 9 cells, 0.5-2000 xlO 6 cells, 0.5-2 xlO 6 cells, 0.5-2 xlO 7 cells, 0.5-2 xlO 8 cells, or 0.5-2 xlO 9 cells, including all integer values within those ranges.
- T cell compositions are administered multiple times at these dosages.
- the dosage is administered a single time or multiple times, for example daily, weekly, biweekly, or monthly, hourly, or is administered upon recurrence, relapse or progression of the cancer being treated.
- the cells in some embodiments, are administered by using infusion techniques that are commonly known in immunotherapy (see, e.g ., Rosenberg et ah, New Eng. J. of Med. 319: 1676, 1988).
- the pharmaceutical composition is substantially free of, e.g. , there are no detectable levels of a contaminant, e.g. , selected from the group consisting of endotoxin, mycoplasma, replication competent lentivirus (RCL), p24, VSV-G nucleic acid, HIV gag, residual anti- CD3/anti-CD28 coated beads, mouse antibodies, pooled human serum, bovine serum albumin, bovine serum, culture media components, vector packaging cell or plasmid components, a bacterium a fungus, mycoplasma, IL-2, and IL-7.
- a contaminant e.g. , selected from the group consisting of endotoxin, mycoplasma, replication competent lentivirus (RCL), p24, VSV-G nucleic acid, HIV gag, residual anti- CD3/anti-CD28 coated beads, mouse antibodies, pooled human serum, bovine serum albumin, bovine serum, culture media components, vector packaging cell or plasmid components, a
- engineered T-cells disclose herein are administered to a subject and blood is subsequently redrawn (or apheresis performed), T-cells therefrom are activated and reinfused into the patient with engineered T cells. This process, in some embodiments, is carried out multiple times every few weeks. T-cells are activated from blood draws of from 10 cc to 400 cc. T-cells are activated from blood draws of 20 cc, 30 cc, 40 cc, 50 cc, 60 cc, 70 cc, 80 cc, 90 cc, or 100 cc.
- the modified/engineered T cells and/or pharmaceutical compositions are administered by methods including, but not limited to, aerosol inhalation, injection, infusion, ingestion, transfusion, implantation or transplantation.
- the modified T cells and/or pharmaceutical compositions are administered to a subject transarterially, subcutaneously, intradermally, intratum orally, intranodally, intramedullary, intramuscularly, by intravenous (i.v.) injection, by intravenous (i.v.) infusion, or intraperitoneally.
- the modified/engineered T cells and/or pharmaceutical compositions thereof are administered to a patient by intradermal or
- modified/engineered T cells and/or pharmaceutical compositions thereof are administered by i.v. injection.
- modified/engineered T cells and/or pharmaceutical compositions thereof are administered by i.v. injection.
- compositions thereof are injected directly into a tumor, lymph node, or site of infection.
- the modified/engineered T cells T cells and/or pharmaceutical compositions are administered in a volume of about 5 mL, 10 mL, 15 mL, 20 mL, 25 mL, 30 mL, 35 mL, 40 mL, 45 mL, 50 mL, 60 mL, 70 mL, 80 mL, 90 mL, 100 mL, 110 mL, 120 mL, 130 mL, 140 mL, 150 mL, 200 mL, 300 mL, 400 mL, or 500 mL.
- the modified/engineered T cells T cells and/or pharmaceutical compositions are administered in a volume of at greater than at most about 5 mL, 10 mL, 15 mL, 20 mL, 25 mL, 30 mL, 35 mL, 40 mL, 45 mL, 50 mL, 60 mL, 70 mL, 80 mL, 90 mL, 100 mL, 110 mL, 120 mL, 130 mL, 140 mL, 150 mL, 200 mL, 300 mL, 400 mL, or 500 mL.
- the modified/engineered T cells T cells and/or pharmaceutical compositions are administered in a volume of at least about 5 mL, 10 mL, 15 mL, 20 mL, 25 mL, 30 mL, 35 mL, 40 mL, 45 mL, 50 mL, 60 mL, 70 mL, 80 mL, 90 mL, 100 mL, 110 mL, 120 mL, 130 mL, 140 mL, 150 mL, 200 mL, 300 mL, 400 mL, or 500 mL.
- a pharmaceutical composition is prepared by per se known methods for the preparation of pharmaceutically acceptable compositions that are administered to subjects, such that an effective quantity of the T cells is combined in a mixture with a pharmaceutically acceptable carrier.
- Suitable carriers are described, for example, in Remington's Pharmaceutical Sciences (Remington's Pharmaceutical Sciences, 20 th ed., Mack Publishing Company, Easton, Pa., USA, 2000).
- the compositions include, albeit not exclusively, solutions of the substances in association with one or more pharmaceutically acceptable carriers or diluents, and contained in buffered solutions with a suitable pH and iso-osmotic with the physiological fluids.
- Suitable pharmaceutically acceptable carriers include essentially chemically inert and nontoxic compositions that do not interfere with the effectiveness of the biological activity of the pharmaceutical composition.
- suitable pharmaceutical carriers include, but are not limited to, water, saline solutions, glycerol solutions, N-(l(2,3-dioleyloxy)propyl)N,N,N- trimethylammonium chloride (DOTMA), diolesylphosphotidyl-ethanolamine (DOPE), and liposomes.
- DOTMA N-(l(2,3-dioleyloxy)propyl)N,N,N- trimethylammonium chloride
- DOPE diolesylphosphotidyl-ethanolamine
- liposomes include a therapeutically effective amount of the compound, together with a suitable amount of carrier so as to provide the form for direct administration to the patient.
- compositions include, without limitation, lyophilized powders or aqueous or non-aqueous sterile injectable solutions or suspensions, which may further contain antioxidants, buffers, bacteriostats and solutes that render the compositions substantially compatible with the tissues or the blood of an intended recipient.
- Other components that may be present in such compositions include water, surfactants (such as Tween), alcohols, polyols, glycerin and vegetable oils, for example.
- Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, tablets, or concentrated solutions or
- a pharmaceutical composition disclosed herein is formulated into a variety of forms and administered by a number of different means.
- a pharmaceutical formulation is administered orally, rectally, or parenterally, in formulations containing conventionally acceptable carriers, adjuvants, and vehicles as desired.
- parenteral as used herein includes subcutaneous, intravenous, intramuscular, or intrastemal injection and infusion techniques.
- Administration includes injection or infusion, including intra-arterial, intracardiac, intracerebroventricular, intradermal, intraduodenal, intramedullary, intramuscular, intraosseous, intraperitoneal, intrathecal, intravascular, intravenous, intravitreal, epidural and subcutaneous), inhalational, transdermal, transmucosal, sublingual, buccal and topical (including epicutaneous, dermal, enema, eye drops, ear drops, intranasal, vaginal) administration.
- intra-arterial intracardiac, intracerebroventricular
- intradermal intraduodenal
- intramedullary intramuscular
- intraosseous intraperitoneal
- intrathecal intravascular, intravenous, intravitreal, epidural and subcutaneous
- inhalational transdermal, transmucosal, sublingual, buccal and topical (including epicutaneous, dermal, enema, eye drops, ear drops, intranasal, vaginal
- a route of administration is via an injection such as an intramuscular, intravenous, subcutaneous, or intraperitoneal injection.
- Liquid formulations include an oral formulation, an intravenous formulation, an intranasal formulation, an ocular formulation, an otic formulation, an aerosol, and the like. In certain embodiments, a combination of various formulations is administered. In certain embodiments a composition is formulated for an extended release profile.
- a target-specific ligand of the TACs disclosed herein bind to a tumor antigen or tumor associated antigen on a tumor cell.
- a target-specific ligand of the TACs disclosed herein selectively bind to a tumor antigen or tumor associated antigen on a tumor cell.
- a target-specific ligand of the TACs disclosed herein specifically bind to a tumor antigen or tumor associated antigen on a tumor cell.
- the target antigen is a tumor antigen.
- tumor antigens include, but are not limited to, CD 19, ITER-2 (erbB-2), B-cell maturation antigen (BCMA), alphafetoprotein (AFP), carcinoembryonic antigen (CEA), CA-125, MUC-l, epithelial tumor antigen (ETA), tyrosinase, melanoma-associated antigen (MAGE), prostate-specific antigen (PSA), glioma-associated antigen, b-human chorionic gonadotropin, thyroglobulin, RAGE-l, MN-CA IX, human telomerase reverse transcriptase, RET1, RET2 (AS), intestinal carboxyl esterase, mut hsp70-2, M-CSF, prostase, PAP, NY-ESO-l, LAGE-la, p53, prostein, PSMA, survivin and telomerase, prostate-carcinoma tumor antigen-l (PCTA-l), ELF2M, neutrophil
- a cancer expressing a target antigen in an individual in need thereof comprising administering to the individual engineered T cells disclosed herein.
- the target antigen is CD 19.
- the method of treating a cancer expressing CD 19 in an individual in need thereof comprises administering to the individual engineered T cells comprising a TAC comprising a CD 19-targeting ligand.
- examples of cancers that are treated by a TAC comprising a CD 19-targeting ligand include, but are not limited to B cell malignancies.
- examples of cancers that are treated by a TAC comprising a CD 19-targeting ligand include, but are not limited to B cell lymphomas, acute lymphoblastic leukemia (ALL), and chronic lymphocytic leukemia (CLL).
- examples of cancers that are treated by a TAC comprising a CD 19-targeting ligand include, but are not limited to Non- Hodgkin’s lymphoma (NHL).
- the target antigen is HER-2.
- the method of treating a cancer wherein a cancer cell expresses HER-2 in an individual in need thereof comprises administering to the individual engineered T cells comprising a TAC comprising a HER-2-targeting ligand.
- examples of cancers that are treated by a TAC comprising a HER-2-targeting ligand include, but are not limited to breast cancer, bladder cancer, pancreatic cancer, ovarian cancer, and stomach cancer.
- the target antigen is BCMA.
- the method of treating a cancer wherein a cancer cell expresses BCMA in an individual in need thereof comprises administering to the individual engineered T cells comprising a TAC comprising a BCMA-targeting ligand.
- examples of cancers that are treated by a TAC comprising a BCMA-targeting ligand include, but are not limited to leukemia, lymphomas, and multiple myeloma.
- an engineered T cell disclosed herein in the preparation of a medicament to treat cancer in an individual in need thereof. Also disclosed herein is the use of a mixture of T cells comprising modified and unmodified cells, or comprising different populations of modified cells with or without unmodified cells.
- a therapeutic quantity of modified T cells need not be homogenous in nature.
- the engineered T cells disclosed herein are part of a combination therapy.
- effectiveness of a therapy disclosure herein is assessed multiple times.
- patients are stratified based on a response to a treatment disclosed herein.
- an effectiveness of treatment determines entrance into a trial.
- cancers that are treated engineered T cells comprising any one of the TAC disclosed herein include any form of neoplastic disease.
- examples of cancers that are treated include, but are not limited to breast cancer, lung cancer and leukemia, for example mixed lineage leukemia (MLL), chronic lymphocytic leukemia (CLL) acute lymphoblastic leukemia (ALL).
- examples of cancers that are treated include, but are not limited to large B-cell lymphoma, diffuse large B-cell lymphoma, primary mediastinal B cell lymphoma, high grade B-cell lymphoma, or large B cell lymphoma arising from follicular lymphoma.
- cancers include carcinomas, blastomas, melanomas, sarcomas, hematological cancers, lymphoid malignancies, benign and malignant tumors, and malignancies.
- the cancer comprises non-solid tumors or solid tumors.
- cancers that are treated include tumors that are not vascularized, or not yet substantially vascularized, as well as vascularized tumors.
- the cancer is a solid cancer or comprises a solid tumor.
- the cancer is a liquid cancer or comprises a liquid tumor.
- the cancer is a lung cancer, a breast cancer, a colon cancer, multiple myeloma, glioblastoma, gastric cancer, ovarian cancer, stomach cancer, colorectal cancer, urothelial cancer, endometrial cancer, or a melanoma.
- the cancer is a lung cancer.
- the cancer is a breast cancer.
- the cancer is a colon cancer.
- the cancer is multiple myeloma.
- the cancer is a glioblastoma.
- the cancer is a gastric cancer.
- the cancer is an ovarian cancer. In some
- the cancer is a stomach cancer. In some embodiments, the cancer is a colorectal cancer. In some embodiments, the cancer is urothelial cancer. In some embodiments, the cancer is an endometrial cancer. In some embodiments, the cancer is a melanoma.
- Fig. 1A shows an example of CD8 T-cell activation based on the co-assembly of different receptors and their associated protein partners.
- the major histocompatibility complex I is presenting an antigen (helix). This is recognized by a T cell receptor (TCR) complex capable of binding the antigen.
- TCR T cell receptor
- the TCR complex contains several individual subunits. The a/b domains are able to interact directly with the antigen presented on MHC-I. The a/b domains then interact with several other domains (e, g, d, and z), all of which participate in T- cell activation via various intracellular activation domains.
- the TCR complex interacts with MHC-I concurrently with the CD8 co-receptor.
- the CD8 co-receptor binds to the MHC-I in an antigen independent manner.
- CD8 directly interacts with Lck, a protein kinase important for activating the TCR receptor complex.
- Lck a protein kinase important for activating the TCR receptor complex.
- the CD8 and Lck interaction also ensures their association with lipid rafts (membrane portion) microdomains, which are hypothesized to organize and encapsulate other relevant signaling moieties (dark spheres). Later stages of activation then lead to CD28 recruitment. If this interaction cascade occurs several times in parallel, T-cells become activated and are able to exert their cytotoxic effects.
- Fig. IB provides an overview of Chimeric Antigen Receptors (CAR).
- CARs seek to reproduce the complex mechanism of T-cell activation by combining several key activation domains, such as O ⁇ 3z and CD28 in a single synthetically engineered molecule. The CAR then directly interacts with an antigen of choice using specific binding domains. Depicted here is an ankyrin repeat protein (DARPin). It is believed that several such interactions occurring in parallel lead to T-cell activation.
- DARPin ankyrin repeat protein
- Fig. 1C is an overview of the Tri-TAC technology mimicking the natural activation process.
- the Tri-TAC was developed to better recapitulate the natural signaling through the TCR, while retaining MHC unrestricted targeting.
- T-cell activation occurs following ligation of MHC by the TCR and T cell co-receptor (either CD4 or CD8), which simultaneously bind to conserved regions within the MHC molecule.
- the co-receptors are specifically located within "lipid rafts", membrane micro domains that are particularly important for TCR signal complex formation.
- these co-receptors In addition to ensuring the correct microdomain localization of the TCR activation complex, these co-receptors also bind directly to Lck, a protein kinase that is crucial for T-cell activation. None of the traditional chimeric receptors or bi-functional proteins engage the co receptor molecules or Lck.
- a molecule was created where the transmembrane and intracellular regions of the CD4 co-receptor, which localize to the lipid raft and bind Lck, respectively, were fused to single-chain antibody that binds CD3 (UCHT1; SEQ ID NO: 13, 14 and homologs thereof). This construct is designed to draw the CD3 molecule and the TCR into regions of lipid rafts and bring Lck into the proximity of the TCR, similar to natural MHC binding.
- Tri-TAC Trifunctional T cell-antigen coupler
- Tri-TAC (Fig. 3A), a Tri-TAC bearing a HER-2-specific DARPin (Fig. 3B), a Tri-TAC bearing a CDl9-specific scFv (Fig. 3C), and a Tri-TAC bearing a BCMA-specific scFv (Fig. 3D).
- Fig. 4A-Fig.4D illustrate the functionality of a Tri-TAC bearing the HER-2-specific DARPin.
- Human T cells were engineered to express either the Tri-TAC as disclosed herein or a conventional CAR with the same DARPin. It was determined that in all aspects, T cells engineered with the Tri-TAC demonstrated functionality at least equivalent to a conventional CAR. Interestingly, with regard to 2 parameters (TNF-a production and CDl07a mobilization), it was observed that the Tri-TAC was more active than a conventional CAR in some
- Fig. 4A shows surface expression of Anti-HER-2 DARPin Tri-TAC compared to Anti-HER-2 DARPin CAR, and control T cells.
- the chimeric receptors were detected by incubation with recombinant HER-2.
- the Anti-HER-2 DARPin Tri-TAC was expressed well on the surface of the engineered T cells.
- Fig. 4B shows growth of the engineered T cells cultures. T cells were activated with anti-CD3/anti-CD28 Dynabeads and engineered with lentiviruses encoding the Tri-TAC, CAR or no receptor (control). After 2 weeks, the CAR and control cultures had grown to similar numbers while the Tri-TAC cultures grew slightly more slowly.
- T cells engineered to express the Tri-TAC or the CAR bearing the HER-2 DARPin were stimulated with plate- bound antigen.
- the T cells engineered to express the Tri-TAC and CAR could elaborate all measured functions (TNF-a production, IFN-g production and CDl07a mobilization, Fig. 3C and Fig. 3D).
- T cells engineered with the Tri-TAC exhibited elevated frequencies of CDl07a- positive cells following stimulation relative to T cells engineered with a CAR (Fig. 3D), suggesting enhanced cytotoxicity on a per-cell basis.
- Fig. 6A-Fig.6J provides data confirming the importance of both ligand binding domain and the ETCHT1 CD3 binding domain for Tri-TAC functionality.
- T cells were engineered with the full-length Tri-TAC bearing the HER-2 DARPin (Fig. 6G, Fig. 6H, Fig. 61, bottom row), a Tri-TAC variant that lacks the DARPin (Fig. 6A, Fig. 6B, Fig. 6C, top row), or a Tri-TAC variant that lacks the UCHT1 (Fig. 6D, Fig. 6E, Fig. 6F, middle row). All three engineered T cell populations were stimulated with HER-2-positive tumor cells.
- T cells engineered with the full-length Tri-TAC could produce IFN-g, TNF- and IL-2 following stimulation, whereas the variants failed to produce any cytokine following stimulation.
- the three T cell populations were also co-cultured with D2F2/E2 cells (HER-2-expressing) or D2F2 cells (HER-2-negative) at an effector: target of 4: 1 (Fig. 6J).
- T cells engineered with full-length Tri-TAC demonstrated robust killing against D2F2/E2 cells but did not kill the D2F2 cells.
- Fig. 7A-Fig. 7C show the results of mice treated with vector control (NGFR), Anti- HER-2 DARPin CAR or Anti-HER-2 DARPin Tri-TAC.
- a xenograft mouse model was used.
- OVCAR-3 tumor cells were administered to mice subcutaneously and allowed to grow until the tumors reached a size of 100 - 200mm 3 .
- Fig. 7 A shows relative tumor progression normalized to tumor size at day of treatment.
- Anti-HER-2 DARPin Tri-TAC engineered T-cells caused a rapid decrease in tumor volume, control had no effect, and CAR cells slowed tumor growth and showed a delayed reduction in tumor size.
- Fig. 7B illustrates relative changes in body weight post T-cell infusion.
- Fig. 7C illustrates cytokine concentrations in serum of mice on day 7 post T-cell infusion. Cytokine levels were higher in CAR-treated mice compared to Tri-TAC -treated mice.
- Fig. 8A-Fig. 8H illustrate the functionality of Tri-TACs bearing alternate CD3 binding domains. The domains are listed in Fig. 8A and Fig. 8E. Tri-TACs containing ETCHT1 (Fig.
- Fig. 8C1 intermediate cytotoxicity
- Fig. 8D intermediate cytotoxicity
- Cells expressing the Tri- TAC containing F6A exhibited strong cytokine production (Fig. 8G, Fig. 8G1) and cytotoxicity (Fig. 8H) following Tri-TAC ligation.
- Cells expressing the Tri-TAC containing L2K exhibited low cytokine production (Fig. 8G, Fig. 8G1) and intermediate cytotoxicity (Fig. 8H).
- Fig. 9A-Fig. 9H illustrates TCR surface expression on T cells engineered with different Tri-TAC variants shown in Fig. 8A and Fig. 8E.
- T cells engineered with the Tri-TAC variants comprising OKT3 (Fig. 9A, Fig. 9E and Fig. 9B, Fig. 9F) or L2K (Fig. 9C, Fig. 9G and Fig 9D, Fig. 9H) exhibited lower TCR surface expression relative to the T cells engineered with Tri- TACs comprising UCHT1 or huUCHTl, respectively.
- T cells engineered with the Tri-TAC variant comprising F6A did not reveal TCR downregulation relative to the Tri-TAC carrying huUCHTl (Fig. 9C, Fig. 9G and Fig. 9D, Fig. 9H).
- the F6A substitution reduced Tri- TAC receptor surface expression, while retaining moderate cytokine production and
- Tri-TAC surface expression and T cell effector functions are not inherently proportional, and that Tri-TAC domain substitutions, in some instances, alters effector functions independent of surface expression levels. It is conceivable that a TAC variant with reduced cytotoxicity and low surface expression could be of value in certain clinical applications.
- the scFv substitutions attenuated the ability of the engineered T cell to elaborate IFN-g, TNF-a, and IL-2, yet the engineered T cells retained the ability to kill target cells.
- Excessive cytokine production has been associated with adverse events in clinical settings, limiting current CAR technologies to life-threatening diseases.
- the ability to modify TAC molecules to reduce their cytokine production while retaining moderate cytotoxicity will allow generation Tri-TAC receptors with the exact level of reactivity required to satisfy clinical efficacy and safety.
- Tri-TAC variant comprising OKT3 to suppress TCR surface expression and cytokine production, while retaining cytotoxicity, could be of great value in allogeneic situations where the suppression of TCR could suppress graft versus host disease.
- Tri-TACs useful in various applications (e.g., oncology, autoimmunity, allergy).
- Fig. IOA-Fig. 10B illustrate several TAC variants with different linkers connecting the ligand that binds a TCR complex and the target-binding ligand domain.
- the flexible connector allows movement between the two domains.
- the large domain connector contains two folded domains and is very large and rigid.
- the small and long helix connectors also introduce rigidity but are less restrictive when compared to the large domain linker.
- Fig. HA-Fig.HE illustrate the impact of connector substitution on Tri-TAC surface expression, Tri-TAC transduction efficiency, and cytokine production upon Tri-TAC ligation.
- Fig. 11A and Fig. 11B show that the helical linkers enhance surface expression and transduction efficiency when compared to the flexible linker, while the large domain connector enhances transduction efficiency but not surface expression.
- Fig. 11D, Fig. HE illustrates cytokine production by cells expressing Tri-TACs with short helix, long helix, or large domain connectors.
- Fig. 12A illustrates enhanced in vitro cytotoxicity of T cells expressing Tri-TACs with the short helix connector.
- Fig. 12B illustrates enhanced in vivo tumor control of T cells expressing Tri-TACs with the short helix connector.
- the short helical connector was associated with high in vitro cytotoxicity and effective in vivo tumor control.
- Fig. 13A illustrates surface expression of CD8a Tri-TAC paired with an anti-HER-2 scFv or Fig. 13C anti-HER-2 DARPin.
- Fig. 13B illustrates cytokine production by T cells expressing CD8a Tri-TAC paired with an anti-HER-2 scFv or anti-HER-2 DARPin.
- Fig. 14A illustrates a CD4 Tri-TAC monomer and a CD8a/p heterodimer.
- TCR co receptors both CD4 and CD8, carry functional domains that are important for the co-receptor functionality. These regions include the arginine rich region that is hypothesized to be important for lipid raft association, and the CXCP motif required for Lck binding.
- ETnlike CD4, which is a monomer the CD8 co-receptor is a heterodimer composed of an a and a b subunit (Fig. 14A). Both the a and b CD8 subunits contain arginine rich regions, but only the a subunit contains the CXCP motif.
- Fig. 14B-Fig. 14D provide schematics of Tri-TAC variants that incorporate elements from the CD8 co-receptor shown in Fig. 14A.
- the cysteine responsible for dimerizing CD8a and O ⁇ 8b was replaced with an alanine in all CD8 Tri-TAC variants.
- Fig. 14B is a schematic of a CD8a Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a CD8a cytosolic domain.
- Fig. 14B is a schematic of a CD8a Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a CD8a cytosolic domain.
- FIG. 14C is a schematic of a CD8aH3 ⁇ 4]3 Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a chimeric CD8a cytosolic domain where the CD8a arginine rich region is replaced with the O ⁇ 8b arginine rich region.
- Fig. 14D is a schematic of a CD8b+Lck Tri-TAC comprising a Cysteine to Serine mutation to ensure a monomeric receptor distribution, and a chimeric O ⁇ 8b cytosolic domain, where the CD8a CXCP domain, which contains an Lck binding motif, was added to the C-terminus of the O ⁇ 8b cytosolic domain.
- Fig. 15A-Fig. 15D illustrate various phenotypic and functional attributes of the CD8- based Tri-TAC variants relative to the prototypical Tri-TAC.
- Fig. 15A-Fig. 15B illustrate surface expression of CD8-Tri TAC variants relative to the prototypic Tri-TAC. Surface expression was comparable among the different Tri-TACs.
- Fig. 15C illustrates in vitro cytotoxicity of CD8-Tri TAC variants co-cultured with LOX IMVI (HER-2 negative) and A549, SKOV3, SKBR3 or MBA MB 231 (all are HER-2 positive). All T cells engineered with Tri- TACs exhibited cytotoxicity.
- Fig. 15D illustrate various phenotypic and functional attributes of the CD8- based Tri-TAC variants relative to the prototypical Tri-TAC.
- Fig. 15A-Fig. 15B illustrate surface expression of CD8-Tri TAC variants relative to the prototypic Tri-TAC. Surface expression was comparable
- FIG. 15D illustrates cell division of T cells engineered with either the CD8 Tri-TAC variants or the prototypic Tri-TAC (Fig. 15D).
- Fig. 15E illustrates TCR surface expression of engineered T cells comprising CD8 Tri-TAC variants or the prototypic Tri-TAC. All Tri-TAC variants had a similar effect on TCR expression. While the CD4 co receptor demonstrated good surface expression and functionality with both the scFv and
- the CD8a construct showed activity only in the context of the DARPin antigen binding domain.
- all the configurations contained the reported key sequence attributes associated with co receptor functionality
- Fig. 16 illustrates the step-wise development of a CD19-TAC construct.
- Several generations of lentiviral vectors are created with various alterations in design elements to ensure CD 19-specificity, proper TAC expression, and GMP-grade lentivirus production.
- Each box represents a lentiviral vector and specifies the 3 major design elements: (A) the antigen -binding domain, (B) the TCR/CD3 -binding domain, and (C) the co-receptor domain. Shaded areas indicate domains that have been the subject of modification during the vector development process.
- the TAC in the first step comprises a HER-2-specific designed ankyrin repeat protein (DARPin), a murine ETCHT1 CD3-specific scFv, and a flexible transmembrane and cytosolic CD4 polypeptide.
- DARPin ankyrin repeat protein
- the TAC is cloned into a pCCL4 lentiviral vector.
- the HER-2-specific DARPin was replaced with a polypeptide comprising an N-terminal CD8a leader peptide fused to an anti-CD 19 scFv.
- the heavy and light chains of the CD 19 scFv were connected via glycine-serine linker region.
- the UCHT1 domain was replaced with a humanized version (huUCHTl) to reduce immunogenicity. This TAC construct exhibited superior surface expression levels than its precursor.
- Fig. 17 illustrates a CD19-TAC insert in a pCCL lentiviral vector.
- the pCCL vector features a bi-directional promoter system with ANGFR(hu) under control of the mCMV promoter and TAC expression being driven by the EF-la promoter.
- the ANGFR(hu) is a truncated human CD271 (Tumor necrosis factor receptor superfamily member 16), with transmembrane domain but lacking the cytosolic signaling domain.
- the ANGFR(hu) expression product is used to quantify lentiviral transduction.
- the CD 19-TAC#921 open reading frame is enlarged to show the key elements of the TAC construct: The CD8a leader, FMC63 single chain (anti-CDl9 scFv), the human c-Myc Tag, the huUCHTl (Y177T) and the ACD4 domain.
- the huUCHTl (Y177T) mutation was identified by examining point mutations randomly introduced into resides of the murine UCHT1 CD3 epsilon binding interface. In a screen the (Y177T) mutation was successfully identified.
- the (Y177T) mutation results in better Tri TAC surface expression while retaining T cell activation.
- ACD4 lacks the four CD4 extracellular
- immunoglobulin like domains retains the extracellular linker, transmembrane and cytosolic domains.
- the CDl9-Tri-TAC construct was cloned into a new lentiviral vector under the control of a MSCV promoter.
- the CDl9-Tri-TAC construct is the same as shown in Fig. 17.
- Example 6 Ability to manufacture CD19-TAC -expressing T cells from different donor material
- Fig. 18 illustrates the efficacy of CD 19 TAC-expressing T cells manufactured from multiple donors.
- CDl9-TAC-expressing T cells were produced using T cells from three different donors, and tested in the NALM-6 tumor model. Mice bearing established NALM-6 tumors were treated with a single dose of 4 x 10 6 CD 19 TAC-expressing T cells. Control mice showed rapid tumor outgrowth, with all mice reaching endpoint by the termination of the study. T cell products from Donors 1 & 2 resulted in complete control in all mice. T cell product from Donor 3 resulted in robust tumor control in all mice and long-term control in 2/4 treated mice. The study confirms that tumor rejection is achieved by CD 19 TAC-expressing T cells derived from multiple healthy donors. The results of the NALM-6 tumor model in Fig. 18 suggest that efficacious CD 19 TAC is produced from multiple donor source materials.
- Example 7 In vitro cytotoxicity and in vivo efficacy of CD19-TAC -expressing T cells
- Tri-TAC-engineered T cells were co-cultured with either NALM-6 (acute lymphoblastic leukemia), Raji (Burkitt lymphoma) or Jeko-l (Mantle Cell Lymphoma).
- NALM-6, Jeko-l and Raji cells were engineered with enhanced firefly luciferase to enable tracking of tumor burden in vitro and in the live animal via bioluminescence imaging.
- Fig. 19A-Fig. 19C illustrates killing of tumor cell lines by CDl9-TAC-expressing T cells.
- the effects were dose-dependent and increased with increasing effector-to-target (E:T) ratios.
- E:T effector-to-target
- As negative controls cells engineered with AT AC (lacking an antigen-binding domain) or non-transduced T cells were used. These results demonstrate that CDl9-TAC-expressing T cells kill CD 19-positive tumor cells.
- Fig. 19 D-Fig. 19G illustrates the design and outcome of an in vivo study assessing efficacy of CD19-TAC in mice engrafted with either NALM-6 (acute lymphoblastic leukemia), Raji (Burkitt lymphoma) or Jeko-l (Mantle Cell Lymphoma) liquid tumors.
- NALM-6 acute lymphoblastic leukemia
- Raji Burkitt lymphoma
- Jeko-l Mantle Cell Lymphoma
- CD ⁇ -TAC-expressing T cells were given as an intravenous tail vein injection. Tumor burden was measured at weekly intervals, and the data are plotted as the average radiance [p/s/cm A 2/sr]
- Fig. 19 E-Fig. 19G illustrates that CD19-TAC engineered T cells are efficacious in inducing tumor regression and long-term tumor control in NALM-6 (acute lymphoblastic leukemia), Raji (Burkitt lymphoma) or Jeko-l (Mantle Cell Lymphoma) liquid tumors.
- NALM-6 acute lymphoblastic leukemia
- Raji Burkitt lymphoma
- Jeko-l Mantle Cell Lymphoma
- Example 8 CD19-TAC -expressing T cell persistence and lasting tumor immunity.
- FIG. 20A- Fig. 20B illustrate persistence of tumor immunity and resistance to re challenge in mice receiving CDl9-TAC-expressing T cells. Mice bearing established NALM-6 tumors were treated with CDl9-TAC-expressing T cells.
- Fig. 20A illustrates the experimental set up to determine CD19-TAC persistence in mice. Mice were first inoculated with NALM-6 cells, which following a 4 day engraftment period were treated with CD19-TAC. All mice showed tumor regression and complete tumor control.
- mice 56 days after the initial treatment mice were re-challenged with either NALM-6 (CD 19 positive) or KMS11 (CD 19 negative) liquid tumors. In all cases naive mice are co-injected with tumor cells and used as negative controls. Tumor burden is followed via luminescence signal.
- Fig. 20B Mice bearing established NALM-6 tumors were treated with CD19-TAC- expressing T cells given as split dose totaling 4 x 10 6 engineered cells. As controls, a group of un-treated animals was used. Following ACT, treated mice presented durable anti-tumor responses. In contrast, control mice showed exponential increases in tumor masses and reached tumor burden related endpoint. On day 56 post-ACT, mice were re-challenged with either NALM-6 tumor cells (CD19 positive) or KMS11 tumor cells (CD19 negative). CD19-TAC- treated mice remain protected from NALM-6 (CD 19 positive) tumor cells, but not from KMS11 (CD 19 negative) tumor cells.
- Example 9 In vivo expansion and dose dependency of CD19-TAC -expressing T cells.
- Fig. 21 and Fig. 22 illustrate dose dependency, dose regime (split or single) and expansion of CD19-TAC -expressing T cells in a NALM-6 cancer model.
- Fig. 21A illustrates experimental design. Mice received either a single dose of CD19-TAC- expressing T cells on day four post-tumor inoculation, or a split dose delivered seven days apart. Multiple CD 19- TAC-expressing T cells doses were tested: 0.5 xlO 6 , 1 xlO 6 , and 4 xlO 6 cells.
- Fig. 21B control groups of mice receive 4 x 10 6 non-transduced cells, or freezing media (vehicle control).
- Fig. 21B illustrate survival of mice after NALM-6 injection and CD19-TAC injection. Dose-dependent promotion of survival were observed both in the single dose and split dose groups, with the highest single administration dose limiting tumor growth and promoting survival the mouse.
- Fig 22A illustrates the gating strategy used to assess T cell proliferation.
- Cells were first selected based on forward and sideways scatter to select for the lymphocyte population. Singlet cells were identified via a forward scatter area over height gate. Live cells were identified via near IR gating. Human cells were identified via a hCD45 gate. The resulting subset of cells was further divided into CD3 positive cells. These cells were then gated on CD4/CD8 and Protein L. The staining strategy also contained muCD45_l to identify murine blood cells. CD 19 was included to stain for NALM-6 cells.
- ACT dose adoptive T cell transfer
- T cells in mice treated with CD19-TAC engineered cells were shown to expand in recipient mice within approximately 1-2 weeks after the first ACT (Fig. 22B). Non-transduced cells did not expand (Fig. 22B).
- Example 10 In vivo efficacy, long term efficacy and safety CD19-TAC treatment
- Fig. 23- Fig. 25 demonstrate the long-term safety and efficacy (Fig. 23) and in the absence of any acute treatment associated toxicities (Fig. 24- Fig. 25).
- Fig. 23A illustrates the experiment design. Mice were injected with 0.5 x 10 6 enhanced luciferase engineered NALM-6 cells, which were allowed to engraft for 4 days. Mice are then treated with two dose levels (4 and 12 x 10 6 engineered cells) of CD 19-T AC-engineered T cells in a single dose administration. Tumor growth was then followed via regular luminesce measurements. Mouse health was regularly assessed via inspection of mouse behavior and physical characteristics (grooming, motility, fur integrity)
- Fig. 23B illustrates the tumor burden via luminescence following treatment with either vehicle alone (Freezing media), non-engineered control cells (Total T cell dose equal to total T cell dose of highest engineered treatment group) and either 4 or 12 x 10 6 engineered CD19-TAC engineered T cells. Both controls show rapid tumor outgrowth and no anti-tumor efficacy. The control dose results in a delay in tumor outgrowth relative to vehicle alone, presumably due to competition between high dose T cell and tumor cells for engraftment niches. Engineered T cell show tumor regression in all cases. High dose treatment groups show complete tumor control in all cases. The 4 x 10 6 treatment group shows 3 mice with complete control, one with delayed tumor outgrowth and one with controlled but high tumor burden.
- Fig. 23C illustrates overall survival of the different treatment groups.
- the vehicle and non-engineered control mice all mice succumb to the tumor within 23 to 35 days respectively.
- all mice develop GvHD symptoms and succumb to GvHD within 61 days.
- GvHD is a consequence of the mouse model itself and not the treatment with the modified T cells.
- Low dose mice show survival of 3 mice until end of study at 90 days, one mouse succumbs to high tumor burden, one mouse succumbs to GvHD.
- Fig. 24 and Fig. 25 illustrates clinical chemistry parameters and cytokine levels from vehicle control, non-engineered and CD19-TAC (4 and 12 x 10 6 effective CD19-TAC engineered cells) treated mice. Mice were followed for 33 days with blood samples taken 5, 12 and 33 days post ACT. Only CD19-TAC treated mice survived for 33 days. Vehicle control mice succumbed to tumor burden before a 3 rd blood sample could be collected, non-engineered cells were sacrificed early on day 26, immediately prior to mice reaching tumor burden related endpoint. All blood samples were analyzed for several clinical chemistry parameters and cytokine levels.
- Fig. 24 illustrates that at day 5 and 12 CD19-TAC treated mice show no parameter that is significantly higher compared to control groups. At day 33 all treated mice show clinical chemistry parameters comparable to early treatment time points, with the exception of Alanine Aminotransferase (ALT) and Aspartate Aminotransferase (AST) where some mice experience high levels, similar to mice treated with non-engineered cells sampled on day 26.
- ALT Alanine Aminotransferase
- AST Aspartate Aminotransferase
- Fig. 25 illustrates the cytokine response on day 5, 12 and 33.
- ACT CD19- TAC but not control mice show elevation in all cytokines tested.
- the cytokine increase is in agreement with an inflammatory response of CD19-TAC engineered T cells recognizing and reacting to antigen positive NALM-6 tumor cells.
- cytokine levels subside, which correlates with the by then induced tumor regression and generally low tumor burden.
- cytokine levels between CD19-TAC treated are either similar or lower than non-engineered T cells except for IL10.
- mice treated with non -transduced or CD19-TAC engineered T cells show an increase in cytokines, presumably associated with GvHD onset. See also Fig. 29, which illustrates cytokine response on day 5, 12, 26 and 33.
- mice treated with CD19-TAC and their clinical chemistry profile demonstrate that engineered T cells are safe to use and do not show any indication of toxicity caused specifically by CD19-TAC engineering.
- the results of the cytokine study demonstrate an early inflammatory response associated with anti-tumor efficacy, following by a drop in all cytokine levels, suggesting that a controlled inflammatory response.
- Fig. 26 illustrates an in vivo efficacy study of various BCMA Tri-TAC constructs.
- Fig. 26A illustrates the overall experimental design. 1 million luciferase-engineered KMS11 (BCMA positive) tumor cells were allowed to engraft for 12 days. Mice were then treated with a single effective dose of 4 million BCMA constructs and controls (Fig. 26B). Tumor burden was regularly assessed via luminescence measurements. All mice that showed tumor regression and tumor control were then re-challenged on day 25 post ACT with 1 million KMS11 cells.
- Fig. 26C Following ACT, control mice exhibited a rapid outgrowth of tumor cells reaching tumor associated endpoint within 19 to 25 days. In contrast all BCMA-TAC treated mice showed initial tumor regression. Tumor control varied across constructs with the G 4 S (SEQ ID NO: 73) 3625VH-VL showing the lowest level of initial tumor control and Short Helix 3625 VL-VH showing the highest level of initial tumor control. Following re-challenge, a majority of all constructs that had maintained tumor control until day 25 remained protected against re challenge.
- G 4 S SEQ ID NO: 73
- mice are inoculated at the hind flank with OVCAR3 solid tumors. Tumors are allowed to establish and grow to a size of lOOmm 3 . Mice are then treated with a tail vain injection of TAC- Her2 engineered T cells. Tumor volume is measured regularly.
- a clinical study is undertaken wherein subjects of at least 18 years of age with CD 19- positive Diffuse Large B-cell Lymphoma who have failed at least two prior lines of therapies including ASCT or who are ineligible for ASCT are treated with CDl9-TAC-expressing T cells.
- the study is an open label, single arm, Phase 1/2 two-stage trial, featuring a dose escalation stage to determine the maximum tolerated dose (MTD) or recommended phase II dose (RPh2D), followed by an expansion cohort at the selected dose.
- MTD maximum tolerated dose
- RPh2D recommended phase II dose
- cyclophosphamide followed by intravenous (IV) administration of CDl9-TAC-expressing T cells.
- IV intravenous
- CDl9-TAC-expressing T cells After treatment with CDl9-TAC-expressing T cells, subjects enter post-treatment follow up and are followed for safety, disease status, and survival for 2 years after their last dose of CDl9-TAC-expressing T cells. After study completion, subjects are followed for survival, long- term toxicity, and viral vector safety in a separate long-term follow up protocol for up to 15 years after their last dose of CDl9-TAC-expressing T cells.
- T cell expansion is assessed from the time of the first dose of CDl9-TAC-expressing T cells until cells are no longer detectable.
- Radiographic disease assessment is performed by positron emission tomography (PET) and/or computed tomography (CT) scans pre-treatment and approximately 3, 6, 9, 12, 18, and 24 months following the last dose of CDl9-TAC-expressing T cells, or until progressive disease, or treatment with additional anti-cancer therapy.
- PET positron emission tomography
- CT computed tomography
- the manufacturing process of CDl9-TAC-expressing T cells drug products involves selecting CD4/CD8 T cells from a leukapheresis product, activating the CD4/CD8 positive cells, transducing the cells with a lentiviral vector comprising the CD19-TAC construct (as described in example 5), expanding the transduced cells to level adequate for the proposed dosing schedule, and harvesting and cryopreserving the final product.
- the patient’s leukapheresis material with its associated unique subject identifier (UPN) is received into a manufacturing site and given a unique specimen number (ISN).
- the CD4/CD8 cells are selected are cryopreserved until initiation of the culture process steps.
- the cryopreserved CD4/CD8 positively selected T cells are thawed at 37°C, resuspended in appropriate medium and seeded into culture bags with activating reagents, the cultures are incubated overnight at 37°C/5% C02.
- the cells are transduced with the CD19-TAC lentiviral vector at an appropriate multiplicity of infection (MOI) and incubated overnight at 37°C/5% C02. On subsequent days, the culture is supplemented with complete medium to maintain a desired cell concentration and eventually pooled into transfer bags, pelleted, resuspended and seeded to larger culture bags at the targeted cell density.
- MOI multiplicity of infection
- the harvested cell suspension is resuspended in excipient and cryopreserved with a controlled-rate freezer then transferred to LN2 storage.
- the product is shipped to the clinical site in its frozen state, thawed at the bedside and administered intravenously.
- Fig. 27 illustrates that TACs proliferate when encountering antigen on cells, but not when the antigen is presented on artificial beads; but CARs proliferate irrespective if antigens are presented on beads or cells.
- Fig. 28A-Fig. 28B illustrate TAC engineered T cells expand in vivo and provide long term protection, indicating cell persistence in a model of myeloma.
- Fig. 28A-Fig. 28B illustrate BCMA-TAC T cells reject multiple myeloma tumors in a KMS-l 1 xenograft model engineered with NanoLuc (KMS 1 l-NanoLuc) (BCMA pos ). Following tumor engraftment mice were treated with BCMA TAC-T cells (carrying Firefly Luciferase). TAC-T cells expand significantly following administration. This correlates with tumor regression. Treated mice were resistant to tumor rechallenge indicating long term persistence of TAC-T cells.
- TAC-T cells destroy tumor cells likely via a mechanism that mimics the natural process of T cell activation.
- the TAC technology illustrates 1) strong efficacy in liquid, 2) in vivo proliferation, 3) T cell persistence, protecting mice from re challenge, and 4) cell expansion following T cell administration.
- Example 16 In vivo and in vitro activity of hu- or mulgic HER2-TAC with MSCV or EFla promoter.
- CD4 and CD8 T cells were engineered with a variety of TAC expressing viruses.
- One set of cells was engineered with a lentivirus that employed the MSCV promoter to express a TAC specific for HER-2 that employed the murine IgG signal peptide [muIgG TAC (MSCV)].
- Another set of cells was engineered with a lentivirus that employed the MSCV promoter to express a TAC specific for HER-2 that employed the human IgG signal peptide [huIgG TAC (MSCV)].
- a third set of cells was engineered with a lentivirus that employed the EFla promoter to express a TAC specific for HER-2 that employed the murine IgG signal peptide [muIgG TAC (EFla)].
- a TAC construct lacking the HER2 binding domain was used as negative control.
- Engineered cells were then characterized in vitro for resurface expression and specific activity and in vivo for activity in the OVCAR3 HER2 -positive solid tumor model.
- Fig. 30 illustrates T cell surface expression of either human or murine IgG leader HER2
- TAC receptors under control of a MCSV or EFla promoter.
- Surface expression is a key requirement for biological activity and this illustrates that the HER2 TAC receptors expression is not influenced by the species source of the IgG signal peptide.
- Fig. 31 demonstrates that T cells engineered with either human or murine IgG leader HER2 TAC constructs, under control of a MCSV or EF la promoter induce cytokine production, when co-cultured with HER2 positive target cells (OVCAR3) but not with HER2 negative cells (LOX IMVI).
- HER2 TAC receptor engineered T cells are capable of specifically engaging HER2 expressing target cells but are non -reactive against antigen negative cells.
- Fig 32 demonstrates the in vivo efficacy of human or murine IgG leader HER2 TAC constructs, under control of a MCSV or EF 1 a promoter. Following split dose administration of engineered T cells, HER2 TAC engineered cells show significant impact on tumor growth, including tumor regression, relative to the negative control Abinding TAC. This in vivo experiment demonstrates that all HER2 TAC engineered cells show significant activity against a solid tumor model in vivo.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Genetics & Genomics (AREA)
- Cell Biology (AREA)
- Biochemistry (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biophysics (AREA)
- Engineering & Computer Science (AREA)
- Epidemiology (AREA)
- Mycology (AREA)
- Oncology (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Toxicology (AREA)
- Gastroenterology & Hepatology (AREA)
- Hematology (AREA)
- General Chemical & Material Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Peptides Or Proteins (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Developmental Biology & Embryology (AREA)
- Virology (AREA)
Abstract
Description
Claims
Priority Applications (16)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
AU2019307905A AU2019307905A1 (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations |
EA202190288A EA202190288A1 (en) | 2019-07-15 | 2019-07-17 | AGENT BINDING T-CELL WITH ANTIGEN WITH DIFFERENT OPTIMIZATION OF CONSTRUCTIONS |
KR1020217004572A KR20210021593A (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various structural optimizations |
CN201980047935.6A CN112689642A (en) | 2018-07-17 | 2019-07-17 | T cell antigen conjugates optimized with various constructs |
EP19838648.4A EP3823984A4 (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations |
JP2020571816A JP7404279B2 (en) | 2018-07-17 | 2019-07-17 | T cell antigen couplers with various construct optimizations |
BR112021000735-0A BR112021000735A2 (en) | 2018-07-17 | 2019-07-17 | t-cell antigen coupler with multiple construct optimizations |
MX2021000541A MX2021000541A (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations. |
CA3104887A CA3104887A1 (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations |
SG11202100307WA SG11202100307WA (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations |
US15/929,513 US11110123B2 (en) | 2018-07-17 | 2020-05-06 | T cell-antigen coupler with various construct optimizations |
IL280049A IL280049A (en) | 2018-07-17 | 2021-01-10 | T cell-antigen coupler with various construct optimizations |
PH12021550114A PH12021550114A1 (en) | 2018-07-17 | 2021-01-15 | T cell-antigen coupler with various construct optimizations |
US17/394,280 US11406667B2 (en) | 2018-07-17 | 2021-08-04 | T cell-antigen coupler with various construct optimizations |
US17/810,238 US20220331364A1 (en) | 2018-07-17 | 2022-06-30 | T cell-antigen coupler with various construct optimizations |
US18/188,326 US11878035B2 (en) | 2018-07-17 | 2023-03-22 | T cell-antigen coupler with various construct optimizations |
Applications Claiming Priority (16)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US201862699173P | 2018-07-17 | 2018-07-17 | |
US62/699,173 | 2018-07-17 | ||
US201862703037P | 2018-07-25 | 2018-07-25 | |
US62/703,037 | 2018-07-25 | ||
US201862773120P | 2018-11-29 | 2018-11-29 | |
US62/773,120 | 2018-11-29 | ||
US201962826853P | 2019-03-29 | 2019-03-29 | |
US62/826,853 | 2019-03-29 | ||
US201962828879P | 2019-04-03 | 2019-04-03 | |
US62/828,879 | 2019-04-03 | ||
US201962839235P | 2019-04-26 | 2019-04-26 | |
US62/839,235 | 2019-04-26 | ||
US16/442,274 US10640562B2 (en) | 2018-07-17 | 2019-06-14 | T cell-antigen coupler with various construct optimizations |
US16/442,274 | 2019-06-14 | ||
US201962874426P | 2019-07-15 | 2019-07-15 | |
US62/874,426 | 2019-07-15 |
Related Parent Applications (2)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
US16/442,274 Continuation-In-Part US10640562B2 (en) | 2018-07-17 | 2019-06-14 | T cell-antigen coupler with various construct optimizations |
US16/442,274 Continuation US10640562B2 (en) | 2018-07-17 | 2019-06-14 | T cell-antigen coupler with various construct optimizations |
Related Child Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
US15/929,513 Continuation US11110123B2 (en) | 2018-07-17 | 2020-05-06 | T cell-antigen coupler with various construct optimizations |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2020018727A1 true WO2020018727A1 (en) | 2020-01-23 |
Family
ID=69163917
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/US2019/042297 WO2020018727A1 (en) | 2018-07-17 | 2019-07-17 | T cell-antigen coupler with various construct optimizations |
Country Status (12)
Country | Link |
---|---|
EP (1) | EP3823984A4 (en) |
JP (1) | JP7404279B2 (en) |
KR (1) | KR20210021593A (en) |
CN (1) | CN112689642A (en) |
AU (1) | AU2019307905A1 (en) |
BR (1) | BR112021000735A2 (en) |
CA (1) | CA3104887A1 (en) |
IL (1) | IL280049A (en) |
MX (1) | MX2021000541A (en) |
PH (1) | PH12021550114A1 (en) |
SG (1) | SG11202100307WA (en) |
WO (1) | WO2020018727A1 (en) |
Cited By (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11001621B1 (en) | 2014-02-07 | 2021-05-11 | Mcmaster University | Trifunctional T cell-antigen coupler and methods and uses thereof |
US11111298B2 (en) | 2018-07-17 | 2021-09-07 | Mcmaster University | T cell-antigen coupler with various construct optimizations |
US11110123B2 (en) | 2018-07-17 | 2021-09-07 | Triumvira Immunologics Usa, Inc. | T cell-antigen coupler with various construct optimizations |
US11198737B2 (en) | 2017-10-12 | 2021-12-14 | Mcmaster University | T cell-antigen coupler with Y177 mutation and methods of uses thereof |
US11453723B1 (en) | 2021-06-25 | 2022-09-27 | Mcmaster University | BCMA T cell-antigen couplers and uses thereof |
WO2022226071A1 (en) | 2021-04-21 | 2022-10-27 | Conagen Inc. | Biosynthetic production of gamma-or delta-lactones using cytochrome p450 hydroxylase enzymes or mutants thereof |
Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2013092001A1 (en) * | 2011-12-19 | 2013-06-27 | Synimmune Gmbh | Bispecific antibody molecule |
US20150322169A1 (en) * | 2012-07-13 | 2015-11-12 | The Trustees Of The University Of Pennsylvania | Enhancing Activity of CAR T Cells by Co-Introducing a Bispecific Antibody |
US20160362472A1 (en) * | 2015-04-08 | 2016-12-15 | Hans Bitter | Cd20 therapies, cd22 therapies, and combination therapies with a cd19 chimeric antigen receptor (car)- expressing cell |
US20160368964A1 (en) * | 2014-02-07 | 2016-12-22 | Mcmaster University | Trifunctional t cell-antigen coupler and methods and uses thereof |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2016166139A1 (en) * | 2015-04-14 | 2016-10-20 | Eberhard Karls Universität Tübingen | Bispecific fusion proteins for enhancing immune responses of lymphocytes against tumor cells |
CN109563507B (en) * | 2016-07-08 | 2024-03-05 | 埃克苏马生物技术公司 | Methods and compositions for transducing lymphocytes and modulating their activity |
EP3493827A4 (en) * | 2016-08-04 | 2020-02-26 | Memorial Sloan-Kettering Cancer Center | Compositions and methods for immunotherapy |
CN108250302A (en) * | 2016-12-29 | 2018-07-06 | 天津天锐生物科技有限公司 | A kind of multifunctional protein |
-
2019
- 2019-07-17 AU AU2019307905A patent/AU2019307905A1/en active Pending
- 2019-07-17 KR KR1020217004572A patent/KR20210021593A/en unknown
- 2019-07-17 JP JP2020571816A patent/JP7404279B2/en active Active
- 2019-07-17 SG SG11202100307WA patent/SG11202100307WA/en unknown
- 2019-07-17 CN CN201980047935.6A patent/CN112689642A/en active Pending
- 2019-07-17 BR BR112021000735-0A patent/BR112021000735A2/en unknown
- 2019-07-17 CA CA3104887A patent/CA3104887A1/en active Pending
- 2019-07-17 EP EP19838648.4A patent/EP3823984A4/en active Pending
- 2019-07-17 WO PCT/US2019/042297 patent/WO2020018727A1/en unknown
- 2019-07-17 MX MX2021000541A patent/MX2021000541A/en unknown
-
2021
- 2021-01-10 IL IL280049A patent/IL280049A/en unknown
- 2021-01-15 PH PH12021550114A patent/PH12021550114A1/en unknown
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2013092001A1 (en) * | 2011-12-19 | 2013-06-27 | Synimmune Gmbh | Bispecific antibody molecule |
US20150322169A1 (en) * | 2012-07-13 | 2015-11-12 | The Trustees Of The University Of Pennsylvania | Enhancing Activity of CAR T Cells by Co-Introducing a Bispecific Antibody |
US20160368964A1 (en) * | 2014-02-07 | 2016-12-22 | Mcmaster University | Trifunctional t cell-antigen coupler and methods and uses thereof |
US20160362472A1 (en) * | 2015-04-08 | 2016-12-15 | Hans Bitter | Cd20 therapies, cd22 therapies, and combination therapies with a cd19 chimeric antigen receptor (car)- expressing cell |
Non-Patent Citations (2)
Title |
---|
ALABANZA, L. ET AL.: "Function of Novel Anti- CD 19 Chimeric Antigen Receptors with Human Variable Regions Is Affected by Hinge and Transmembrane Domains", MOLECULAR THERAPY, vol. 25, no. 11, 2017, pages 2452 - 2465, XP055505801, DOI: 10.1016/j.ymthe.2017.07.013 * |
See also references of EP3823984A4 * |
Cited By (15)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11421014B2 (en) | 2014-02-07 | 2022-08-23 | Mcmaster University | Trifunctional T cell-antigen coupler and methods and uses thereof |
US11008376B2 (en) | 2014-02-07 | 2021-05-18 | Mcmaster University | Trifunctional T cell-antigen coupler and methods and uses thereof |
US11001621B1 (en) | 2014-02-07 | 2021-05-11 | Mcmaster University | Trifunctional T cell-antigen coupler and methods and uses thereof |
US11970545B2 (en) | 2017-10-12 | 2024-04-30 | Mcmaster University | T cell-antigen coupler with Y182T mutation and methods of uses thereof |
US11198737B2 (en) | 2017-10-12 | 2021-12-14 | Mcmaster University | T cell-antigen coupler with Y177 mutation and methods of uses thereof |
US11643472B2 (en) | 2017-10-12 | 2023-05-09 | Mcmaster University | T cell-antigen coupler with Y182T mutation and methods and uses thereof |
US11110123B2 (en) | 2018-07-17 | 2021-09-07 | Triumvira Immunologics Usa, Inc. | T cell-antigen coupler with various construct optimizations |
US11406667B2 (en) | 2018-07-17 | 2022-08-09 | Triumvira Immunologies USA, Inc. | T cell-antigen coupler with various construct optimizations |
US11878035B2 (en) | 2018-07-17 | 2024-01-23 | Triumvira Immunologics Usa, Inc. | T cell-antigen coupler with various construct optimizations |
US11111298B2 (en) | 2018-07-17 | 2021-09-07 | Mcmaster University | T cell-antigen coupler with various construct optimizations |
US11976117B2 (en) | 2018-07-17 | 2024-05-07 | Mcmaster University | T cell-antigen coupler with various construct optimizations |
WO2022226071A1 (en) | 2021-04-21 | 2022-10-27 | Conagen Inc. | Biosynthetic production of gamma-or delta-lactones using cytochrome p450 hydroxylase enzymes or mutants thereof |
WO2022226065A2 (en) | 2021-04-21 | 2022-10-27 | Conagen Inc. | Biosynthetic production of delta-lactones using cytochrome p450 hydroxylase enzymes or mutants thereof |
US11453723B1 (en) | 2021-06-25 | 2022-09-27 | Mcmaster University | BCMA T cell-antigen couplers and uses thereof |
WO2022266778A1 (en) * | 2021-06-25 | 2022-12-29 | Mcmaster University | Bcma t cell-antigen couplers and uses thereof |
Also Published As
Publication number | Publication date |
---|---|
JP2021530971A (en) | 2021-11-18 |
EP3823984A1 (en) | 2021-05-26 |
KR20210021593A (en) | 2021-02-26 |
IL280049A (en) | 2021-03-01 |
BR112021000735A2 (en) | 2021-07-13 |
JP7404279B2 (en) | 2023-12-25 |
CA3104887A1 (en) | 2020-01-23 |
SG11202100307WA (en) | 2021-02-25 |
MX2021000541A (en) | 2021-08-16 |
CN112689642A (en) | 2021-04-20 |
EP3823984A4 (en) | 2022-04-06 |
PH12021550114A1 (en) | 2021-10-04 |
AU2019307905A1 (en) | 2021-03-04 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
US11111298B2 (en) | T cell-antigen coupler with various construct optimizations | |
US11878035B2 (en) | T cell-antigen coupler with various construct optimizations | |
US11421014B2 (en) | Trifunctional T cell-antigen coupler and methods and uses thereof | |
JP7404279B2 (en) | T cell antigen couplers with various construct optimizations | |
US11970545B2 (en) | T cell-antigen coupler with Y182T mutation and methods of uses thereof | |
US20230364237A1 (en) | Claudin 18.2 t cell-antigen couplers and uses thereof | |
EP4240381A1 (en) | Cells comprising t cell-antigen couplers and uses thereof | |
EA045368B1 (en) | T-CELL ANTIGEN BINDING AGENT WITH VARIOUS OPTIMIZATION OF DESIGNS | |
WO2023283111A2 (en) | Gucy2c t cell-antigen couplers and uses thereof | |
EP4367133A2 (en) | Gucy2c t cell-antigen couplers and uses thereof |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 19838648 Country of ref document: EP Kind code of ref document: A1 |
|
ENP | Entry into the national phase |
Ref document number: 2020571816 Country of ref document: JP Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 3104887 Country of ref document: CA |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
REG | Reference to national code |
Ref country code: BR Ref legal event code: B01A Ref document number: 112021000735 Country of ref document: BR |
|
ENP | Entry into the national phase |
Ref document number: 20217004572 Country of ref document: KR Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 2019838648 Country of ref document: EP Effective date: 20210217 |
|
ENP | Entry into the national phase |
Ref document number: 2019307905 Country of ref document: AU Date of ref document: 20190717 Kind code of ref document: A |
|
REG | Reference to national code |
Ref country code: BR Ref legal event code: B01E Ref document number: 112021000735 Country of ref document: BR Free format text: APRESENTE O COMPLEMENTO DO TEXTO EM PORTUGUES, ADAPTADO A NORMA VIGENTE, DO PEDIDO CONFORME DEPOSITO INTERNACIONAL INICIAL (RELATORIO DESCRITIVO), CONFORME DETERMINAA RESOLUCAO INPI PR NO 77/2013 DE 18/03/2013, ART. 5O E 7O. |
|
REG | Reference to national code |
Ref country code: BR Ref legal event code: B01E Ref document number: 112021000735 Country of ref document: BR Free format text: APRESENTE O COMPLEMENTO DO TEXTO EM PORTUGUES, ADAPTADO A NORMA VIGENTE, DO PEDIDO CONFORME DEPOSITO INTERNACIONAL INICIAL (RELATORIO DESCRITIVO), CONFORME DETERMINAA RESOLUCAO INPI PR NO 77/2013 DE 18/03/2013, ART. 5O E 7O. |
|
REG | Reference to national code |
Ref country code: BR Ref legal event code: B01Y Ref document number: 112021000735 Country of ref document: BR Free format text: ANULADA A PUBLICACAO CODIGO 1.5 NA RPI NO 2625 DE 27/04/2021 POR TER SIDO INDEVIDA. |
|
ENP | Entry into the national phase |
Ref document number: 112021000735 Country of ref document: BR Kind code of ref document: A2 Effective date: 20210115 |