CN117715649A - Methods of treating Acute Respiratory Distress Syndrome (ARDS) in a particular patient using mesenchymal lineage precursor or stem cells - Google Patents
Methods of treating Acute Respiratory Distress Syndrome (ARDS) in a particular patient using mesenchymal lineage precursor or stem cells Download PDFInfo
- Publication number
- CN117715649A CN117715649A CN202280034316.5A CN202280034316A CN117715649A CN 117715649 A CN117715649 A CN 117715649A CN 202280034316 A CN202280034316 A CN 202280034316A CN 117715649 A CN117715649 A CN 117715649A
- Authority
- CN
- China
- Prior art keywords
- subject
- cells
- mlpsc
- stem cells
- ards
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 239000002243 precursor Substances 0.000 title claims abstract description 151
- 210000000130 stem cell Anatomy 0.000 title claims abstract description 147
- 238000000034 method Methods 0.000 title claims abstract description 136
- 206010001052 Acute respiratory distress syndrome Diseases 0.000 title claims abstract description 95
- 239000000203 mixture Substances 0.000 claims abstract description 78
- 210000004027 cell Anatomy 0.000 claims description 234
- 239000000090 biomarker Substances 0.000 claims description 160
- 230000002757 inflammatory effect Effects 0.000 claims description 149
- 208000013616 Respiratory Distress Syndrome Diseases 0.000 claims description 74
- 201000000028 adult respiratory distress syndrome Diseases 0.000 claims description 74
- 238000011282 treatment Methods 0.000 claims description 70
- 206010061218 Inflammation Diseases 0.000 claims description 41
- 230000004054 inflammatory process Effects 0.000 claims description 41
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 41
- 230000002829 reductive effect Effects 0.000 claims description 41
- 210000004072 lung Anatomy 0.000 claims description 37
- 210000000440 neutrophil Anatomy 0.000 claims description 37
- 102000019034 Chemokines Human genes 0.000 claims description 36
- 108010012236 Chemokines Proteins 0.000 claims description 36
- 210000002540 macrophage Anatomy 0.000 claims description 36
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 35
- 230000037396 body weight Effects 0.000 claims description 34
- 102000008857 Ferritin Human genes 0.000 claims description 28
- 108050000784 Ferritin Proteins 0.000 claims description 28
- 238000008416 Ferritin Methods 0.000 claims description 28
- 101710187743 Tumor necrosis factor receptor superfamily member 1A Proteins 0.000 claims description 25
- 102100033732 Tumor necrosis factor receptor superfamily member 1A Human genes 0.000 claims description 25
- 230000004202 respiratory function Effects 0.000 claims description 24
- 239000003246 corticosteroid Substances 0.000 claims description 22
- 102100021943 C-C motif chemokine 2 Human genes 0.000 claims description 21
- 102100025248 C-X-C motif chemokine 10 Human genes 0.000 claims description 20
- 101000858088 Homo sapiens C-X-C motif chemokine 10 Proteins 0.000 claims description 20
- 102100032366 C-C motif chemokine 7 Human genes 0.000 claims description 19
- 101000797758 Homo sapiens C-C motif chemokine 7 Proteins 0.000 claims description 19
- 108091006905 Human Serum Albumin Proteins 0.000 claims description 19
- 102000008100 Human Serum Albumin Human genes 0.000 claims description 19
- 108090001007 Interleukin-8 Proteins 0.000 claims description 19
- 108090001005 Interleukin-6 Proteins 0.000 claims description 18
- 102100036170 C-X-C motif chemokine 9 Human genes 0.000 claims description 17
- 101000897480 Homo sapiens C-C motif chemokine 2 Proteins 0.000 claims description 17
- 101000947172 Homo sapiens C-X-C motif chemokine 9 Proteins 0.000 claims description 17
- 230000002459 sustained effect Effects 0.000 claims description 17
- 102000004889 Interleukin-6 Human genes 0.000 claims description 16
- 230000009467 reduction Effects 0.000 claims description 16
- 102100036842 C-C motif chemokine 19 Human genes 0.000 claims description 15
- 101000713106 Homo sapiens C-C motif chemokine 19 Proteins 0.000 claims description 15
- 108010002350 Interleukin-2 Proteins 0.000 claims description 15
- 230000004941 influx Effects 0.000 claims description 14
- 102000000588 Interleukin-2 Human genes 0.000 claims description 13
- 230000005012 migration Effects 0.000 claims description 13
- 238000013508 migration Methods 0.000 claims description 13
- 241000711573 Coronaviridae Species 0.000 claims description 12
- 102100028990 C-X-C chemokine receptor type 3 Human genes 0.000 claims description 11
- 101000916050 Homo sapiens C-X-C chemokine receptor type 3 Proteins 0.000 claims description 11
- 230000006044 T cell activation Effects 0.000 claims description 11
- 230000006052 T cell proliferation Effects 0.000 claims description 11
- 230000006907 apoptotic process Effects 0.000 claims description 11
- 239000003527 fibrinolytic agent Substances 0.000 claims description 11
- 208000036142 Viral infection Diseases 0.000 claims description 10
- 230000004913 activation Effects 0.000 claims description 10
- 229960000103 thrombolytic agent Drugs 0.000 claims description 10
- 230000009385 viral infection Effects 0.000 claims description 10
- 230000000735 allogeneic effect Effects 0.000 claims description 9
- 239000003443 antiviral agent Substances 0.000 claims description 9
- 241000725643 Respiratory syncytial virus Species 0.000 claims description 8
- 230000003247 decreasing effect Effects 0.000 claims description 8
- 102100031151 C-C chemokine receptor type 2 Human genes 0.000 claims description 7
- 101710149815 C-C chemokine receptor type 2 Proteins 0.000 claims description 7
- 241000315672 SARS coronavirus Species 0.000 claims description 7
- 210000001744 T-lymphocyte Anatomy 0.000 claims description 6
- FVVLHONNBARESJ-NTOWJWGLSA-H magnesium;potassium;trisodium;(2r,3s,4r,5r)-2,3,4,5,6-pentahydroxyhexanoate;acetate;tetrachloride;nonahydrate Chemical compound O.O.O.O.O.O.O.O.O.[Na+].[Na+].[Na+].[Mg+2].[Cl-].[Cl-].[Cl-].[Cl-].[K+].CC([O-])=O.OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C([O-])=O FVVLHONNBARESJ-NTOWJWGLSA-H 0.000 claims description 6
- 241000127282 Middle East respiratory syndrome-related coronavirus Species 0.000 claims description 5
- 229960003957 dexamethasone Drugs 0.000 claims description 5
- UREBDLICKHMUKA-CXSFZGCWSA-N dexamethasone Chemical compound C1CC2=CC(=O)C=C[C@]2(C)[C@]2(F)[C@@H]1[C@@H]1C[C@@H](C)[C@@](C(=O)CO)(O)[C@@]1(C)C[C@@H]2O UREBDLICKHMUKA-CXSFZGCWSA-N 0.000 claims description 5
- 241000709661 Enterovirus Species 0.000 claims description 4
- 239000000872 buffer Substances 0.000 claims description 4
- 241000712461 unidentified influenza virus Species 0.000 claims description 4
- -1 CCL3 Proteins 0.000 claims description 3
- 230000001419 dependent effect Effects 0.000 claims description 2
- 102000004890 Interleukin-8 Human genes 0.000 claims 1
- 102100032752 C-reactive protein Human genes 0.000 description 47
- 239000000523 sample Substances 0.000 description 43
- 230000014509 gene expression Effects 0.000 description 25
- 230000008859 change Effects 0.000 description 23
- 238000002659 cell therapy Methods 0.000 description 20
- 108090000623 proteins and genes Proteins 0.000 description 20
- 238000005138 cryopreservation Methods 0.000 description 19
- 239000003550 marker Substances 0.000 description 19
- 239000000243 solution Substances 0.000 description 19
- 102100026236 Interleukin-8 Human genes 0.000 description 17
- 229940096397 interleukin-8 Drugs 0.000 description 16
- XKTZWUACRZHVAN-VADRZIEHSA-N interleukin-8 Chemical compound C([C@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@@H](NC(C)=O)CCSC)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(=O)N[C@@H](CCSC)C(=O)N1[C@H](CCC1)C(=O)N1[C@H](CCC1)C(=O)N[C@@H](C)C(=O)N[C@H](CC(O)=O)C(=O)N[C@H](CCC(O)=O)C(=O)N[C@H](CC(O)=O)C(=O)N[C@H](CC=1C=CC(O)=CC=1)C(=O)N[C@H](CO)C(=O)N1[C@H](CCC1)C(N)=O)C1=CC=CC=C1 XKTZWUACRZHVAN-VADRZIEHSA-N 0.000 description 16
- 102100031092 C-C motif chemokine 3 Human genes 0.000 description 15
- 101000777387 Homo sapiens C-C motif chemokine 3 Proteins 0.000 description 15
- 229940100601 interleukin-6 Drugs 0.000 description 15
- 239000002609 medium Substances 0.000 description 14
- 238000003556 assay Methods 0.000 description 13
- 230000006872 improvement Effects 0.000 description 13
- 208000007536 Thrombosis Diseases 0.000 description 12
- 102000004169 proteins and genes Human genes 0.000 description 12
- 210000004369 blood Anatomy 0.000 description 11
- 239000008280 blood Substances 0.000 description 11
- 230000037361 pathway Effects 0.000 description 11
- 238000004458 analytical method Methods 0.000 description 10
- 230000034994 death Effects 0.000 description 10
- 230000002085 persistent effect Effects 0.000 description 10
- 210000001519 tissue Anatomy 0.000 description 10
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 9
- 239000000543 intermediate Substances 0.000 description 9
- 238000002360 preparation method Methods 0.000 description 9
- 239000003795 chemical substances by application Substances 0.000 description 8
- 201000010099 disease Diseases 0.000 description 8
- 102100025683 Alkaline phosphatase, tissue-nonspecific isozyme Human genes 0.000 description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 7
- 239000000546 pharmaceutical excipient Substances 0.000 description 7
- JLTPSDHKZGWXTD-UHFFFAOYSA-N 2-[6-(dicyanomethylidene)naphthalen-2-ylidene]propanedinitrile Chemical compound N#CC(C#N)=C1C=CC2=CC(=C(C#N)C#N)C=CC2=C1 JLTPSDHKZGWXTD-UHFFFAOYSA-N 0.000 description 6
- 101710161969 Alkaline phosphatase, tissue-nonspecific isozyme Proteins 0.000 description 6
- 208000025721 COVID-19 Diseases 0.000 description 6
- 102000003810 Interleukin-18 Human genes 0.000 description 6
- 108090000171 Interleukin-18 Proteins 0.000 description 6
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 6
- 230000000840 anti-viral effect Effects 0.000 description 6
- 210000000988 bone and bone Anatomy 0.000 description 6
- 238000003018 immunoassay Methods 0.000 description 6
- 108020004707 nucleic acids Proteins 0.000 description 6
- 102000039446 nucleic acids Human genes 0.000 description 6
- 150000007523 nucleic acids Chemical class 0.000 description 6
- 238000006213 oxygenation reaction Methods 0.000 description 6
- 208000024891 symptom Diseases 0.000 description 6
- 102000003390 tumor necrosis factor Human genes 0.000 description 6
- 102000004127 Cytokines Human genes 0.000 description 5
- 108090000695 Cytokines Proteins 0.000 description 5
- 108010017213 Granulocyte-Macrophage Colony-Stimulating Factor Proteins 0.000 description 5
- 102100039620 Granulocyte-macrophage colony-stimulating factor Human genes 0.000 description 5
- 102000003816 Interleukin-13 Human genes 0.000 description 5
- 108090000176 Interleukin-13 Proteins 0.000 description 5
- 102000004388 Interleukin-4 Human genes 0.000 description 5
- 108090000978 Interleukin-4 Proteins 0.000 description 5
- 108010023197 Streptokinase Proteins 0.000 description 5
- 239000000427 antigen Substances 0.000 description 5
- 108091007433 antigens Proteins 0.000 description 5
- 102000036639 antigens Human genes 0.000 description 5
- 210000001185 bone marrow Anatomy 0.000 description 5
- 229960001334 corticosteroids Drugs 0.000 description 5
- 238000012258 culturing Methods 0.000 description 5
- 230000006378 damage Effects 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 239000001963 growth medium Substances 0.000 description 5
- 229940028885 interleukin-4 Drugs 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 230000004048 modification Effects 0.000 description 5
- 238000012986 modification Methods 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 210000002966 serum Anatomy 0.000 description 5
- 230000004083 survival effect Effects 0.000 description 5
- 238000002560 therapeutic procedure Methods 0.000 description 5
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 4
- 230000001464 adherent effect Effects 0.000 description 4
- 239000006143 cell culture medium Substances 0.000 description 4
- 230000010261 cell growth Effects 0.000 description 4
- 210000002950 fibroblast Anatomy 0.000 description 4
- 238000007710 freezing Methods 0.000 description 4
- 230000008014 freezing Effects 0.000 description 4
- 238000000338 in vitro Methods 0.000 description 4
- 230000002401 inhibitory effect Effects 0.000 description 4
- 238000011221 initial treatment Methods 0.000 description 4
- 238000003752 polymerase chain reaction Methods 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 238000010257 thawing Methods 0.000 description 4
- 206010003178 Arterial thrombosis Diseases 0.000 description 3
- 206010067671 Disease complication Diseases 0.000 description 3
- 206010061818 Disease progression Diseases 0.000 description 3
- 208000000059 Dyspnea Diseases 0.000 description 3
- 206010013975 Dyspnoeas Diseases 0.000 description 3
- 238000002965 ELISA Methods 0.000 description 3
- 102100037241 Endoglin Human genes 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 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 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 101000881679 Homo sapiens Endoglin Proteins 0.000 description 3
- 101000738771 Homo sapiens Receptor-type tyrosine-protein phosphatase C Proteins 0.000 description 3
- 229920001612 Hydroxyethyl starch Polymers 0.000 description 3
- 108010064136 Monocyte Chemoattractant Proteins Proteins 0.000 description 3
- 102000014962 Monocyte Chemoattractant Proteins Human genes 0.000 description 3
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- 208000010378 Pulmonary Embolism Diseases 0.000 description 3
- 102100037422 Receptor-type tyrosine-protein phosphatase C Human genes 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 210000001789 adipocyte Anatomy 0.000 description 3
- 230000003321 amplification Effects 0.000 description 3
- 210000004556 brain Anatomy 0.000 description 3
- 239000000969 carrier Substances 0.000 description 3
- 239000013592 cell lysate Substances 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 230000000875 corresponding effect Effects 0.000 description 3
- 239000002577 cryoprotective agent Substances 0.000 description 3
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical group P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 3
- 230000005750 disease progression Effects 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 210000003494 hepatocyte Anatomy 0.000 description 3
- 229940050526 hydroxyethylstarch Drugs 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 208000015181 infectious disease Diseases 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- 210000003205 muscle Anatomy 0.000 description 3
- 238000003199 nucleic acid amplification method Methods 0.000 description 3
- 210000003668 pericyte Anatomy 0.000 description 3
- 239000002953 phosphate buffered saline Substances 0.000 description 3
- 108090000765 processed proteins & peptides Proteins 0.000 description 3
- 230000004044 response Effects 0.000 description 3
- 108010051412 reteplase Proteins 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 238000003860 storage Methods 0.000 description 3
- 210000002435 tendon Anatomy 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- 230000003442 weekly effect Effects 0.000 description 3
- VBICKXHEKHSIBG-UHFFFAOYSA-N 1-monostearoylglycerol Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCC(O)CO VBICKXHEKHSIBG-UHFFFAOYSA-N 0.000 description 2
- 102100022464 5'-nucleotidase Human genes 0.000 description 2
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 2
- 108010058207 Anistreplase Proteins 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- 239000003154 D dimer Substances 0.000 description 2
- 101000678236 Homo sapiens 5'-nucleotidase Proteins 0.000 description 2
- 101000599951 Homo sapiens Insulin-like growth factor I Proteins 0.000 description 2
- 101000800116 Homo sapiens Thy-1 membrane glycoprotein Proteins 0.000 description 2
- 108010034143 Inflammasomes Proteins 0.000 description 2
- 102100037852 Insulin-like growth factor I Human genes 0.000 description 2
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 2
- 102000043131 MHC class II family Human genes 0.000 description 2
- 108091054438 MHC class II family Proteins 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- 108700018351 Major Histocompatibility Complex Proteins 0.000 description 2
- 108010010416 Myogenic Regulatory Factors Proteins 0.000 description 2
- 102000015864 Myogenic Regulatory Factors Human genes 0.000 description 2
- 102100038380 Myogenic factor 5 Human genes 0.000 description 2
- 208000037847 SARS-CoV-2-infection Diseases 0.000 description 2
- 201000003176 Severe Acute Respiratory Syndrome Diseases 0.000 description 2
- 241000700584 Simplexvirus Species 0.000 description 2
- 102100033523 Thy-1 membrane glycoprotein Human genes 0.000 description 2
- 108010000134 Vascular Cell Adhesion Molecule-1 Proteins 0.000 description 2
- 102100023543 Vascular cell adhesion protein 1 Human genes 0.000 description 2
- 206010047249 Venous thrombosis Diseases 0.000 description 2
- 241000700605 Viruses Species 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 229960001997 adefovir Drugs 0.000 description 2
- WOZSCQDILHKSGG-UHFFFAOYSA-N adefovir depivoxil Chemical compound N1=CN=C2N(CCOCP(=O)(OCOC(=O)C(C)(C)C)OCOC(=O)C(C)(C)C)C=NC2=C1N WOZSCQDILHKSGG-UHFFFAOYSA-N 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 230000000890 antigenic effect Effects 0.000 description 2
- 210000001130 astrocyte Anatomy 0.000 description 2
- 239000011324 bead Substances 0.000 description 2
- 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 2
- 230000004071 biological effect Effects 0.000 description 2
- 210000004271 bone marrow stromal cell Anatomy 0.000 description 2
- 210000000845 cartilage Anatomy 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 210000001612 chondrocyte Anatomy 0.000 description 2
- 230000003021 clonogenic effect Effects 0.000 description 2
- 210000002808 connective tissue Anatomy 0.000 description 2
- 231100000135 cytotoxicity Toxicity 0.000 description 2
- 230000003013 cytotoxicity Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- 239000003937 drug carrier Substances 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 210000002919 epithelial cell Anatomy 0.000 description 2
- 210000004700 fetal blood Anatomy 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- 108010052295 fibrin fragment D Proteins 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 230000012010 growth Effects 0.000 description 2
- 239000003102 growth factor Substances 0.000 description 2
- 210000002216 heart Anatomy 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 208000014674 injury Diseases 0.000 description 2
- 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 2
- 229910052742 iron Inorganic materials 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 239000008101 lactose Substances 0.000 description 2
- 210000003041 ligament Anatomy 0.000 description 2
- 239000003446 ligand Substances 0.000 description 2
- 210000004185 liver Anatomy 0.000 description 2
- 210000004698 lymphocyte Anatomy 0.000 description 2
- 238000007403 mPCR Methods 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000013028 medium composition Substances 0.000 description 2
- 210000003716 mesoderm Anatomy 0.000 description 2
- 239000002207 metabolite Substances 0.000 description 2
- 210000004165 myocardium Anatomy 0.000 description 2
- 230000001537 neural effect Effects 0.000 description 2
- 210000004498 neuroglial cell Anatomy 0.000 description 2
- 210000002569 neuron Anatomy 0.000 description 2
- 238000010899 nucleation Methods 0.000 description 2
- 210000004248 oligodendroglia Anatomy 0.000 description 2
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 2
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 2
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 2
- 238000004393 prognosis Methods 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 102000005962 receptors Human genes 0.000 description 2
- 108020003175 receptors Proteins 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- 229960002917 reteplase Drugs 0.000 description 2
- 208000013220 shortness of breath Diseases 0.000 description 2
- 210000002027 skeletal muscle Anatomy 0.000 description 2
- 210000002363 skeletal muscle cell Anatomy 0.000 description 2
- 210000003491 skin Anatomy 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 229960005202 streptokinase Drugs 0.000 description 2
- 210000002536 stromal cell Anatomy 0.000 description 2
- 230000020382 suppression by virus of host antigen processing and presentation of peptide antigen via MHC class I Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000011287 therapeutic dose Methods 0.000 description 2
- 230000035899 viability Effects 0.000 description 2
- 206010047470 viral myocarditis Diseases 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 1
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 description 1
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 1
- AEQDJSLRWYMAQI-UHFFFAOYSA-N 2,3,9,10-tetramethoxy-6,8,13,13a-tetrahydro-5H-isoquinolino[2,1-b]isoquinoline Chemical compound C1CN2CC(C(=C(OC)C=C3)OC)=C3CC2C2=C1C=C(OC)C(OC)=C2 AEQDJSLRWYMAQI-UHFFFAOYSA-N 0.000 description 1
- BDKLKNJTMLIAFE-UHFFFAOYSA-N 2-(3-fluorophenyl)-1,3-oxazole-4-carbaldehyde Chemical compound FC1=CC=CC(C=2OC=C(C=O)N=2)=C1 BDKLKNJTMLIAFE-UHFFFAOYSA-N 0.000 description 1
- VHRSUDSXCMQTMA-PJHHCJLFSA-N 6alpha-methylprednisolone Chemical compound C([C@@]12C)=CC(=O)C=C1[C@@H](C)C[C@@H]1[C@@H]2[C@@H](O)C[C@]2(C)[C@@](O)(C(=O)CO)CC[C@H]21 VHRSUDSXCMQTMA-PJHHCJLFSA-N 0.000 description 1
- 241000502561 Acacia irrorata Species 0.000 description 1
- 102100034594 Angiopoietin-1 Human genes 0.000 description 1
- 210000002237 B-cell of pancreatic islet Anatomy 0.000 description 1
- 102000004506 Blood Proteins Human genes 0.000 description 1
- 108010017384 Blood Proteins Proteins 0.000 description 1
- 108010074051 C-Reactive Protein Proteins 0.000 description 1
- 101150013553 CD40 gene Proteins 0.000 description 1
- 102100032912 CD44 antigen Human genes 0.000 description 1
- 108010084313 CD58 Antigens Proteins 0.000 description 1
- 206010008479 Chest Pain Diseases 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 102000029816 Collagenase Human genes 0.000 description 1
- 108060005980 Collagenase Proteins 0.000 description 1
- 208000001528 Coronaviridae Infections Diseases 0.000 description 1
- 206010011224 Cough Diseases 0.000 description 1
- 206010011703 Cyanosis Diseases 0.000 description 1
- 101100239693 Dictyostelium discoideum myoD gene Proteins 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
- 208000005189 Embolism Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102100037362 Fibronectin Human genes 0.000 description 1
- 108010067306 Fibronectins Proteins 0.000 description 1
- 102000003688 G-Protein-Coupled Receptors Human genes 0.000 description 1
- 108090000045 G-Protein-Coupled Receptors Proteins 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 102000053187 Glucuronidase Human genes 0.000 description 1
- 108010060309 Glucuronidase Proteins 0.000 description 1
- 108010017080 Granulocyte Colony-Stimulating Factor Proteins 0.000 description 1
- 102000004269 Granulocyte Colony-Stimulating Factor Human genes 0.000 description 1
- 102100032510 Heat shock protein HSP 90-beta Human genes 0.000 description 1
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 description 1
- 108700005087 Homeobox Genes Proteins 0.000 description 1
- 101000574445 Homo sapiens Alkaline phosphatase, tissue-nonspecific isozyme Proteins 0.000 description 1
- 101000924552 Homo sapiens Angiopoietin-1 Proteins 0.000 description 1
- 101000868273 Homo sapiens CD44 antigen Proteins 0.000 description 1
- 101001016856 Homo sapiens Heat shock protein HSP 90-beta Proteins 0.000 description 1
- 101000998178 Homo sapiens Interleukin-17C Proteins 0.000 description 1
- 101001055222 Homo sapiens Interleukin-8 Proteins 0.000 description 1
- 101000692455 Homo sapiens Platelet-derived growth factor receptor beta Proteins 0.000 description 1
- 101000914514 Homo sapiens T-cell-specific surface glycoprotein CD28 Proteins 0.000 description 1
- 101000914484 Homo sapiens T-lymphocyte activation antigen CD80 Proteins 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 102100034343 Integrase Human genes 0.000 description 1
- 108010061833 Integrases Proteins 0.000 description 1
- 108010064593 Intercellular Adhesion Molecule-1 Proteins 0.000 description 1
- 102100037877 Intercellular adhesion molecule 1 Human genes 0.000 description 1
- 102100033105 Interleukin-17C Human genes 0.000 description 1
- 208000022120 Jeavons syndrome Diseases 0.000 description 1
- 102000007547 Laminin Human genes 0.000 description 1
- 108010085895 Laminin Proteins 0.000 description 1
- 101000988090 Leishmania donovani Heat shock protein 83 Proteins 0.000 description 1
- 108060001084 Luciferase Proteins 0.000 description 1
- 239000005089 Luciferase Substances 0.000 description 1
- 208000030289 Lymphoproliferative disease Diseases 0.000 description 1
- 102000043129 MHC class I family Human genes 0.000 description 1
- 108091054437 MHC class I family Proteins 0.000 description 1
- 208000001145 Metabolic Syndrome Diseases 0.000 description 1
- 108700011259 MicroRNAs Proteins 0.000 description 1
- 101100013973 Mus musculus Gata4 gene Proteins 0.000 description 1
- 206010028391 Musculoskeletal Pain Diseases 0.000 description 1
- 208000009525 Myocarditis Diseases 0.000 description 1
- 108010020197 Myogenic Regulatory Factor 5 Proteins 0.000 description 1
- 101710099061 Myogenic factor 5 Proteins 0.000 description 1
- 108010056785 Myogenin Proteins 0.000 description 1
- 102000004364 Myogenin Human genes 0.000 description 1
- GXCLVBGFBYZDAG-UHFFFAOYSA-N N-[2-(1H-indol-3-yl)ethyl]-N-methylprop-2-en-1-amine Chemical compound CN(CCC1=CNC2=C1C=CC=C2)CC=C GXCLVBGFBYZDAG-UHFFFAOYSA-N 0.000 description 1
- 229930193140 Neomycin Natural products 0.000 description 1
- 102000014413 Neuregulin Human genes 0.000 description 1
- 108050003475 Neuregulin Proteins 0.000 description 1
- 102000048238 Neuregulin-1 Human genes 0.000 description 1
- 108090000556 Neuregulin-1 Proteins 0.000 description 1
- 101800000675 Neuregulin-2 Proteins 0.000 description 1
- 101800000673 Neuregulin-3 Proteins 0.000 description 1
- 206010067482 No adverse event Diseases 0.000 description 1
- 208000008589 Obesity Diseases 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 208000006735 Periostitis Diseases 0.000 description 1
- 102100026547 Platelet-derived growth factor receptor beta Human genes 0.000 description 1
- 206010035664 Pneumonia Diseases 0.000 description 1
- 229920002732 Polyanhydride Polymers 0.000 description 1
- 229920000954 Polyglycolide Polymers 0.000 description 1
- 229920001710 Polyorthoester Polymers 0.000 description 1
- 108010076039 Polyproteins Proteins 0.000 description 1
- 102100022668 Pro-neuregulin-2, membrane-bound isoform Human genes 0.000 description 1
- 102100022659 Pro-neuregulin-3, membrane-bound isoform Human genes 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 108010029485 Protein Isoforms Proteins 0.000 description 1
- 102000001708 Protein Isoforms Human genes 0.000 description 1
- 108700008625 Reporter Genes Proteins 0.000 description 1
- 208000004756 Respiratory Insufficiency Diseases 0.000 description 1
- 208000037656 Respiratory Sounds Diseases 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 229920002472 Starch Polymers 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
- 102100027213 T-cell-specific surface glycoprotein CD28 Human genes 0.000 description 1
- 102100027222 T-lymphocyte activation antigen CD80 Human genes 0.000 description 1
- 101150052863 THY1 gene Proteins 0.000 description 1
- GYDJEQRTZSCIOI-UHFFFAOYSA-N Tranexamic acid Chemical compound NCC1CCC(C(O)=O)CC1 GYDJEQRTZSCIOI-UHFFFAOYSA-N 0.000 description 1
- 108091023040 Transcription factor Proteins 0.000 description 1
- 102000040945 Transcription factor Human genes 0.000 description 1
- 102000004887 Transforming Growth Factor beta Human genes 0.000 description 1
- 108090001012 Transforming Growth Factor beta Proteins 0.000 description 1
- 102400001320 Transforming growth factor alpha Human genes 0.000 description 1
- 101800004564 Transforming growth factor alpha Proteins 0.000 description 1
- 102000008579 Transposases Human genes 0.000 description 1
- 108010020764 Transposases Proteins 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 102100040245 Tumor necrosis factor receptor superfamily member 5 Human genes 0.000 description 1
- 206010047472 Viral pericarditis Diseases 0.000 description 1
- 206010047924 Wheezing Diseases 0.000 description 1
- 201000000690 abdominal obesity-metabolic syndrome Diseases 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 208000038016 acute inflammation Diseases 0.000 description 1
- 230000006022 acute inflammation Effects 0.000 description 1
- 238000004115 adherent culture Methods 0.000 description 1
- 210000000577 adipose tissue Anatomy 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 239000003146 anticoagulant agent Substances 0.000 description 1
- 239000004599 antimicrobial Substances 0.000 description 1
- 210000001367 artery Anatomy 0.000 description 1
- 210000000013 bile duct Anatomy 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 108091006374 cAMP receptor proteins Proteins 0.000 description 1
- 210000004413 cardiac myocyte Anatomy 0.000 description 1
- 238000012832 cell culture technique Methods 0.000 description 1
- 230000003915 cell function Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 239000002458 cell surface marker Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000004087 circulation Effects 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 229960002424 collagenase Drugs 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 230000001086 cytosolic effect Effects 0.000 description 1
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 1
- 238000002716 delivery method Methods 0.000 description 1
- 210000004443 dendritic cell Anatomy 0.000 description 1
- 210000003074 dental pulp Anatomy 0.000 description 1
- 210000004207 dermis Anatomy 0.000 description 1
- 239000008121 dextrose Substances 0.000 description 1
- 239000000539 dimer Substances 0.000 description 1
- 208000035475 disorder Diseases 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 230000003828 downregulation Effects 0.000 description 1
- 210000003981 ectoderm Anatomy 0.000 description 1
- 239000012636 effector Substances 0.000 description 1
- 210000001162 elastic cartilage Anatomy 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 210000001900 endoderm Anatomy 0.000 description 1
- 230000003511 endothelial effect Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 210000000981 epithelium Anatomy 0.000 description 1
- 230000007717 exclusion Effects 0.000 description 1
- 210000001808 exosome Anatomy 0.000 description 1
- 206010016256 fatigue Diseases 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 235000013312 flour Nutrition 0.000 description 1
- 238000000684 flow cytometry Methods 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 238000012239 gene modification Methods 0.000 description 1
- 230000005017 genetic modification Effects 0.000 description 1
- 235000013617 genetically modified food Nutrition 0.000 description 1
- 210000004602 germ cell Anatomy 0.000 description 1
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 1
- YQEMORVAKMFKLG-UHFFFAOYSA-N glycerine monostearate Natural products CCCCCCCCCCCCCCCCCC(=O)OC(CO)CO YQEMORVAKMFKLG-UHFFFAOYSA-N 0.000 description 1
- SVUQHVRAGMNPLW-UHFFFAOYSA-N glycerol monostearate Natural products CCCCCCCCCCCCCCCCC(=O)OCC(O)CO SVUQHVRAGMNPLW-UHFFFAOYSA-N 0.000 description 1
- 150000002334 glycols Chemical class 0.000 description 1
- 210000003780 hair follicle Anatomy 0.000 description 1
- 210000000442 hair follicle cell Anatomy 0.000 description 1
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 1
- 230000003258 hemophagocytic effect Effects 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- 229960002897 heparin Drugs 0.000 description 1
- 229920002674 hyaluronan Polymers 0.000 description 1
- 229960003160 hyaluronic acid Drugs 0.000 description 1
- 210000004408 hybridoma Anatomy 0.000 description 1
- 208000037850 immune anergy Diseases 0.000 description 1
- 230000016784 immunoglobulin production Effects 0.000 description 1
- 238000003364 immunohistochemistry Methods 0.000 description 1
- 239000003547 immunosorbent Substances 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 208000027866 inflammatory disease Diseases 0.000 description 1
- 206010022000 influenza Diseases 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 230000035992 intercellular communication Effects 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000000644 isotonic solution Substances 0.000 description 1
- 210000002510 keratinocyte Anatomy 0.000 description 1
- 101150066555 lacZ gene Proteins 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 230000004199 lung function Effects 0.000 description 1
- 210000001165 lymph node Anatomy 0.000 description 1
- 201000001268 lymphoproliferative syndrome Diseases 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 210000004962 mammalian cell Anatomy 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 238000005399 mechanical ventilation Methods 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 229960004584 methylprednisolone Drugs 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 210000003097 mucus Anatomy 0.000 description 1
- 208000010125 myocardial infarction Diseases 0.000 description 1
- 230000001114 myogenic effect Effects 0.000 description 1
- 229960004927 neomycin Drugs 0.000 description 1
- 210000003061 neural cell Anatomy 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 235000020824 obesity Nutrition 0.000 description 1
- 210000004416 odontoblast Anatomy 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 210000003463 organelle Anatomy 0.000 description 1
- 210000000963 osteoblast Anatomy 0.000 description 1
- 210000000496 pancreas Anatomy 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000009054 pathological process Effects 0.000 description 1
- 208000008494 pericarditis Diseases 0.000 description 1
- 210000003460 periosteum Anatomy 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
- 230000002688 persistence Effects 0.000 description 1
- 238000000053 physical method Methods 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 239000013612 plasmid Substances 0.000 description 1
- 229920000747 poly(lactic acid) Polymers 0.000 description 1
- 239000004633 polyglycolic acid Substances 0.000 description 1
- 239000004626 polylactic acid Substances 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 229960004618 prednisone Drugs 0.000 description 1
- XOFYZVNMUHMLCC-ZPOLXVRWSA-N prednisone Chemical compound O=C1C=C[C@]2(C)[C@H]3C(=O)C[C@](C)([C@@](CC4)(O)C(=O)CO)[C@@H]4[C@@H]3CCC2=C1 XOFYZVNMUHMLCC-ZPOLXVRWSA-N 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 210000001147 pulmonary artery Anatomy 0.000 description 1
- 230000003134 recirculating effect Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 201000004193 respiratory failure Diseases 0.000 description 1
- 230000029058 respiratory gaseous exchange Effects 0.000 description 1
- 208000023504 respiratory system disease Diseases 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 229940116243 retavase Drugs 0.000 description 1
- 210000001525 retina Anatomy 0.000 description 1
- 210000000844 retinal pigment epithelial cell Anatomy 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000012679 serum free medium Substances 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 210000004927 skin cell Anatomy 0.000 description 1
- 210000002460 smooth muscle Anatomy 0.000 description 1
- 210000000329 smooth muscle myocyte Anatomy 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 229940087562 sodium acetate trihydrate Drugs 0.000 description 1
- 239000000176 sodium gluconate Substances 0.000 description 1
- 235000012207 sodium gluconate Nutrition 0.000 description 1
- 229940005574 sodium gluconate Drugs 0.000 description 1
- RYYKJJJTJZKILX-UHFFFAOYSA-M sodium octadecanoate Chemical compound [Na+].CCCCCCCCCCCCCCCCCC([O-])=O RYYKJJJTJZKILX-UHFFFAOYSA-M 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 238000009168 stem cell therapy Methods 0.000 description 1
- 238000009580 stem-cell therapy Methods 0.000 description 1
- 238000012105 stratification Analysis Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 229920002994 synthetic fiber Polymers 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- 230000002381 testicular Effects 0.000 description 1
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 230000004797 therapeutic response Effects 0.000 description 1
- 230000002537 thrombolytic effect Effects 0.000 description 1
- 210000001541 thymus gland Anatomy 0.000 description 1
- 238000010361 transduction Methods 0.000 description 1
- 230000026683 transduction Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
- 238000011269 treatment regimen Methods 0.000 description 1
- 230000001960 triggered effect Effects 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 210000004926 tubular epithelial cell Anatomy 0.000 description 1
- 230000003827 upregulation Effects 0.000 description 1
- 210000003556 vascular endothelial cell Anatomy 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 238000009423 ventilation Methods 0.000 description 1
- 239000013603 viral vector Substances 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
- 230000004580 weight loss Effects 0.000 description 1
- 208000016261 weight loss Diseases 0.000 description 1
Landscapes
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The present disclosure relates to methods for treating or preventing Acute Respiratory Distress Syndrome (ARDS) in a subject in need thereof, the methods comprising administering to the subject a composition comprising mesenchymal lineage precursor or stem cells (MLPSCs).
Description
Technical Field
The present disclosure relates to methods for treating or preventing Acute Respiratory Distress Syndrome (ARDS) in a subject in need thereof.
Background
Respiratory diseases associated with a variety of disorders, such as viral infections, are a problem for the general population. In many cases, they are accompanied by inflammation, which exacerbates lung conditions and may lead to Acute Respiratory Distress Syndrome (ARDS). Patients over 65 are more likely to have increased disease severity and worse clinical outcome than patients under 65.
There remains an unmet need for treatment in elderly patients with Acute Respiratory Distress Syndrome (ARDS), particularly when secondary to viral infection and new treatment regimens are needed.
Disclosure of Invention
The inventors have demonstrated that administration of mesenchymal lineage or precursor cells (MLPSC) can improve respiratory function in patients suffering from Acute Respiratory Distress Syndrome (ARDS). Biomarker analysis revealed down-regulation of inflammatory pathways in patients < 65 years of age treated with MLPSCs, and this corresponds to improved prognosis and persistent improvement in respiratory function. Corresponding biomarker analysis of MLPSC treated patients older than 65 years surprisingly revealed that these patients had higher baseline levels of inflammation, and this corresponds to initial improvement in respiratory function, but did not persist. Since similar inflammatory pathways are involved in ARDS patients regardless of age (< 65 years relative to > 65 years), the inventors' findings indicate that improved treatment of patients with increased baseline levels of inflammatory biomarkers and/or > 65 years can be achieved if patients are treated with higher doses of MLPSC or longer doses. In certain instances, higher doses of MLPSC or longer administrations can achieve results comparable to those of < 65 years old patients.
Thus, in a first example, the present disclosure is directed to a method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising selecting a subject with ARDS greater than or equal to 65 years of age and administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg).
In one example, the subject's inflammatory biomarker level is elevated, which indicates:
increased flow of neutrophils and macrophages into the lung;
-increased macrophage inflammation and increased neutrophil migration to the lung; and/or the number of the groups of groups,
t cell activation/proliferation and apoptotic death.
In another example, the present disclosure relates to a method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising selecting a subject with ARDS having an elevated baseline inflammation level. In one example, an increase in baseline inflammation level is determined based on the increased baseline inflammation level. Thus, in one example, the present disclosure relates to a method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising selecting a subject with an ARDS having an increased inflammatory biomarker indicative of:
increased flow of neutrophils and macrophages into the lung;
-increased macrophage inflammation and increased neutrophil migration to the lung; and/or the number of the groups of groups,
t cell activation/proliferation and apoptotic death,
and administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg).
In another example, the disclosure relates to a method of treating or preventing Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg), wherein the subject is greater than or equal to 65 years old and/or has an increased inflammatory biomarker indicative of:
increased flow of neutrophils and macrophages into the lung;
-increased macrophage inflammation and increased neutrophil migration to the lung; and/or the number of the groups of groups,
t cell activation/proliferation and apoptotic death.
In one example, the subject is ventilator dependent. For example, the subject can be mechanically ventilated prior to administration of the MLPSC.
In one example, the one or more inflammatory biomarkers that indicate increased neutrophil and macrophage influx into the lung are CCR2 or CXCR3 binding chemokines. For example, the CCR2 binding chemokine may be CCL2, CCL3, or CCL7. In one example, the CCR2 binding chemokine is CCL2. In another example, the CXCR3 binding chemokine is CXCL10 or CXCL9. In one example, the CXCR3 binding chemokine is CXCL10. In one example, the one or more inflammatory biomarkers indicative of increased macrophage inflammation and increased neutrophil migration to the lung is IL-6 or IL-8. In one example, the one or more inflammatory biomarkers indicative of T cell activation/proliferation and apoptotic death is CCL19 or IL-2.
The findings of the present inventors indicate that subjects with elevated levels of inflammatory biomarkers can be selected for effective treatment based on the level and/or age of certain or certain biomarkers. In one example, the methods of the present disclosure include selecting a subject based on an increase in CCL2 levels relative to a subject < 65 years old. In another example, subjects are selected based on increased levels of CCL3 relative to subjects < 65 years old. In another example, subjects are selected based on increased levels of CCL7 relative to subjects < 65 years old. In another example, the methods of the present disclosure include selecting a subject based on an increase in CXCL10 levels relative to a subject < 65 years old. In another example, subjects are selected based on an increase in CXCL9 levels relative to subjects < 65 years old. In another example, the methods of the present disclosure include selecting a subject based on an increase in IL-6 levels relative to a subject < 65 years old. In another example, the subject is selected based on an increase in IL-8 levels relative to a subject < 65 years old. In another example, the methods of the present disclosure comprise selecting a subject based on an increase in CCL19 levels relative to a subject < 65 years old. In another example, the subject is selected based on an increase in IL-2 levels relative to a subject < 65 years old. In these examples, the level of inflammatory biomarkers is increased by more than a factor of 1 relative to a patient < 65 years old. In another example, the level of inflammatory biomarker is increased by at least 2-fold relative to a patient < 65 years old. In another example, the level of the inflammatory biomarker is increased by at least a factor of 3 relative to a patient < 65 years old. In another example, the level of the inflammatory biomarker is increased by at least 4-fold relative to a patient < 65 years old. In another example, the level of the inflammatory biomarker is increased by at least 5-fold relative to a patient < 65 years old. In another example, the level of inflammatory biomarker is increased by more than 5-fold relative to a patient < 65 years old. In one example, patients less than 65 years old are not taking corticosteroids.
In another example, the subject is selected based on sustained CRP levels after administration of the first dose of MLPSC. In one example, subjects are selected based on sustained CRP levels 7 days after administration of the first dose of MLPSC. In one example, subjects are selected based on sustained CRP levels 14 days after administration of the first dose of MLPSC. In one example, subjects are selected based on sustained CRP levels 21 days after administration of the first dose of MLPSC. In one example, subjects are selected based on sustained CRP levels 30 days after administration of the first dose of MLPSC.
In one example, treatment with the methods disclosed herein allows the subject to leave the ventilator. In one example, the subject is out of the ventilator within 60 days of treatment.
In one example, the present disclosure relates to a method of treating or preventing Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising: selecting a subject, the subject:
-greater than or equal to 65 years old; and/or the number of the groups of groups,
-an elevated level of one or more inflammatory biomarkers relative to a subject less than 65 years old; and/or the number of the groups of groups,
-having sustained CRP levels 3 days after administration of a first dose of mesenchymal lineage precursor or stem cells (MLPSC) comprising less than 400 ten thousand MLPSCs per kilogram body weight (cells/kg); and
Over 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg) are administered to the subject. In one example, the method further comprises determining or having determined the subject level of one or more inflammatory biomarkers selected from the list comprising: (i) CXCR3 binds to chemokines, preferably CXCL10 and/or CXCL9; (ii) CCR 2-binding chemokines, preferably CCL2, CCL3 and/or CCL7; (iii) IL-6; (v) IL-8; (vi) CCL19; (vii) IL-2; and/or (viii) CRP.
The inventors have also identified that treatment with cell therapy can reduce the level of inflammatory biomarkers in a patient. Thus, in another example, treatment with a composition according to the present disclosure reduces the level of at least one inflammatory biomarker, which indicates:
-reduced flow of neutrophils and macrophages into the lungs;
-reduction of the inflammasome;
-reduced macrophage activation and neutrophil migration to the lung;
-reduced T cell influx and activation; or (b)
-circulating biomarkers for reducing macrophage and neutrophil inflammation.
In one example, the inflammatory biomarker is a CXCR 3-binding chemokine, such as CXCL10. In another example, the CXCR 3-binding chemokine is CXCL9. In another example, the inflammatory biomarker is CCR 2-binding chemokine, such as CCL2. In another example, the CCR 2-binding chemokine is CCL3. In another example, the CCR 2-binding chemokine is CCL7. In another example, the inflammatory biomarker is IL-6. In another example, the inflammatory biomarker is IL-8. In another example, the inflammatory biomarker is TNF. In another example, the inflammatory biomarker is IL-18. In another example, the inflammatory biomarker is CCL19. In another example, the inflammatory biomarker is IL-4. In another example, the inflammatory biomarker is IL-13. In another example, the inflammatory biomarker is GM-CSF. In another example, the inflammatory biomarker is CRP. In another example, the inflammatory biomarker is ferritin.
In one example, treatment according to the present disclosure reduces CRP and/or ferritin levels within 3 to 14 days of administration of more than 400 ten thousand MLPSCs per kilogram body weight (cells/kg). In another example, the treatment improves respiratory function in the subject. For example, berlin standards may be used to determine improved respiratory function. In one example, the treatment improves the berlin standard of the subject on day 14 and/or day 21. In one example, the improved respiratory function is maintained after day 7 relative to the baseline respiratory function. In one example, the improved respiratory function is maintained relative to the baseline respiratory function at day 14.
In one example, more than 500 ten thousand MLPSCs/kg are administered to a subject. In another example, more than 600 ten thousand MLPSCs/kg are administered to a subject. In another example, more than 800 ten thousand MLPSCs/kg are administered to a subject. In one example, more than 400 ten thousand MLPSCs/kg are administered by at least 2 to 3 doses. In one example, over 400 ten thousand MLPSCs/kg are administered by 3 doses. In one example, the subject receives more than 400 ten thousand MLPSCs per kilogram body weight (cells per kilogram) within 5 to 9 days of administration of the first dose of MLPSCs. In one example, the methods of the present disclosure include administering 1 x 10 8 Up to 2.5X10 8 Amount between MLPSC/dose. In another example, the methods of the present disclosure include administering about 1.6x10 8 MLPSC/dose. In another example, the methods of the present disclosure include administering about 200 ten thousand MLPSCs/kg/dose.
In one example, the ARDS of the subject is moderate or severe.
In one example, the subject is taking a corticosteroid prior to administration of the cell composition disclosed herein. In another example, a method according to the present disclosure further comprises administering a corticosteroid. In one example, the corticosteroid is dexamethasone.
In one example, ARDS is caused by a viral infection. Viral infections may be caused by, for example, rhinoviruses, influenza viruses, respiratory Syncytial Viruses (RSV) or coronaviruses.
In one example, ARDS is caused by coronavirus infection. The coronavirus may be, for example, severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV), COVID-19, 229E, NL, OC43 or KHU1. In one example, the coronavirus is SARS-CoV, MERS-CoV or COVID-19 (SARS-CoV-2). In one example, ARDS is secondary to SARS-CoV-2 infection.
In one example, ARDS is caused by thrombosis, such as venous thrombosis or arterial thrombosis. In another example, ARDS is caused by pulmonary embolism.
In one example, the MLPSC has been cryopreserved and thawed. In one example, MLPSCs are culture expanded from an intermediate cryopreserved population of MLPSCs. In another example, the MLPSC culture is amplified for at least about 5 generations. In one example, the MLPSC expresses at least 13pg TNF-R1/million MLPSCs. In one example, the MLPSC expresses about 13pg to about 44pg TNF-R1/million MLPSCs. In one example, culture-amplified MLPSCs are culture-amplified at least 20 population doublings. In another example, culture-amplified MLPSCs are culture-amplified at least 30 population doublings. In one example, the MLPSC is a Mesenchymal Stem Cell (MSC). In another example, the MLPSC is allogeneic. For example, the MLPSC may be an allogeneic MSC.
In another example, the MLPSC is modified to carry or express an antiviral drug or thrombolytic agent. In one example, the antiviral drug is adefovir. In one example, the thrombolytic agent is selected from the group consisting of: eminase (anipulase), retase (reteplase), streppase (streptokinase), and kabikinase (kabikinase).
In another example, the MLPSC is genetically modified to express an antiviral peptide or nucleic acid encoding the same.
In one example, the composition is administered intravenously.
In one example, more than 400 ten thousand MLPSCs/kg are administered to a subject by at least 2 to 3 doses. In one example, the subject receives more than 400 ten thousand MLPSCs/kg within 5 to 9 days of administration of the first dose. In another example, the subject receives more than 400 ten thousand MLPSCs/kg within 7 days of administration of the first dose. In these examples, the dose is contained in 1X 10 8 And 2.5X10 8 Amount between individual cells/dose. For example, the dosage comprises 1.6X10 8 Individual cells. In another example, the dose comprises 200 tens of thousands of cells/kg.
In one example, the MLPSCs of the present disclosure are administered in a composition. In one example, the composition further comprises Plasma-Lyte a, dimethyl sulfoxide (DMSO), human Serum Albumin (HSA). In one example, the composition further comprises a solution of P1asma-Lyte a (70%), DMSO (10%), HSA (25%), the HSA solution comprising 5% HSA and 15% buffer.
In one example, the composition comprises more than 6.68X10 6 Each living cell/mL.
In another example, the present disclosure relates to a method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising:
-administering 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg) to the subject;
-determining or having determined the subject level of one or more biomarkers selected from the group consisting of: CXCL10, CXCL9, CCL2, CCL3, CCL7, IL-6, IL-8, CCL19, IL-2, ferritin and/or CRP;
if the level of one or more biomarkers does not decrease from baseline, then an additional dose of MLPSC is administered to the subject, wherein after the additional dose, more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg) have been administered to the subject.
In one example, the biomarker is CRP and/or ferritin. In one example, CRP and/or ferritin levels are determined for the subject relative to a baseline on day 7 and/or day 14.
In one example, another dose comprises 200 ten thousand MLPSCs per kilogram body weight (cells/kg).
Drawings
Fig. 1: on day 60, cell therapy provided protection from death in patients < 65 years old. A: ITT patients < 65 years old; b: ITT patients are more than or equal to 65 years old.
Fig. 2: on day 60, cell therapy provided protection from death in patients < 65 years old. A: PP patient < 65 years (n=123); b: PP patients were > 65 years old (n=94).
Fig. 3: ARRDS severity (as measured by regression and/or improvement of ARDS as defined by the Berlin standard) in patients < 65 years and patients > 65 years.
Fig. 4: a: CRP levels at baseline and at days 3, 7 and 14; b: ferritin levels at baseline and at days 3, 7 and 14; c: d-dimer at baseline and at days 3, 7 and 14.
Fig. 5: patients with age > 65 had higher levels of baseline inflammation. The data are fold changes in level.
Fig. 6: inflammatory biomarker stratification analysis by age group. Data are fold changes in level from baseline.
Detailed Description
Throughout this specification, unless the context clearly dictates otherwise, reference to a single step, composition of matter, group of steps or group of compositions of matter should be taken to encompass both one step, composition of matter, group of steps or group of compositions of matter as well as multiple steps (i.e., one or more).
It will be appreciated by those skilled in the art that the disclosure described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the present disclosure includes all such variations and modifications. The disclosure also includes all of the steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations or any two or more of said steps or features.
The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended as illustrations only. Functionally equivalent products, compositions, and methods, as described herein, are clearly within the scope of the disclosure.
Any examples disclosed herein should be considered as applicable to any other examples mutatis mutandis unless explicitly stated otherwise.
Unless specifically defined otherwise, all technical and scientific terms used herein shall be taken to have the same meaning as commonly understood by one of ordinary skill in the art (e.g., in cell culture, molecular genetics, stem cell therapy, immunology, immunohistochemistry, protein chemistry, biochemistry).
Unless otherwise indicated, surgical techniques used in this disclosure are standard procedures well known to those skilled in the art.
Methods for obtaining and enriching mesenchymal lineage stem cells or precursor cell populations are known in the art. For example, an enriched population of mesenchymal lineage stem cells or precursor cells can be obtained by using flow cytometry and cell sorting procedures based on the use of cell surface markers expressed on mesenchymal lineage stem cells or precursor cells.
All documents cited or referenced herein, and all documents cited or referenced in the documents cited herein, are hereby incorporated by reference in their entireties, along with any manufacturer's instructions, descriptions, product specifications, and product sheets for any product mentioned herein or in any document incorporated by reference.
Selected definition
The term "and/or", e.g. "X and/or Y", is understood to mean "X and Y" or "X or Y", and should be used to provide explicit support for both meanings or for either meaning.
As used herein, the term "about," unless stated to the contrary, means +/-10%, more preferably +/-5%, of the specified value.
The terms "level" and "amount" are used to define the amount of a particular substance in a sample or cell preparation (or sample thereof) from a subject. For example, a particular concentration, weight, percentage (e.g., v/v%) or ratio may be used to define the level of a particular substance.
In one example, the level of a particular marker is determined in one or more samples obtained from a patient or subject (e.g., a blood sample, a plasma sample, or a serum sample). For example, the level of an inflammatory biomarker according to the present disclosure may be determined in a blood sample.
In one example, the level of an inflammatory biomarker is determined by measuring the level of protein expression in a sample obtained from a subject. There are a variety of assays known in the art that can be used to measure the protein expression level of an inflammatory biomarker in a sample. For example, the inflammatory biomarker levels may be measured in the sample using an antibody-based immunoassay, such as an enzyme-linked immunosorbent assay (ELISA). In one example, a blood sample is obtained from a patient and then purified prior to contact with an antibody that binds to an inflammatory biomarker according to the present disclosure. In this example, the extent of antibody binding is used to quantify the level of inflammatory biomarker (e.g., pg/mL) in the sample. In another example, inflammatory biomarker levels can be measured in a sample using a multiplex immunoassay, e.g., a Luminex assay (see, e.g., cook et al methods 158:27-32.2019). In another example, a fluorescent bead-based immunoassay can be used to measure inflammatory biomarker levels in a sample. In these examples, protein expression of multiple inflammatory biomarkers is measured in a single sample. In another example, protein expression of an inflammatory biomarker is measured in a separate sample.
In another example, the level of an inflammatory biomarker is determined by measuring the level of gene expression in a sample obtained from a subject. There are a variety of assays known in the art that can be used to measure the level of gene expression of an inflammatory biomarker in a sample. For example, a molecular-based assay, such as a qualitative Polymerase Chain Reaction (PCR) based assay, may be used to measure inflammatory biomarker levels in a sample. In one example, a blood sample is obtained from a subject and then purified and lysed to obtain a blood cell lysate. In this example, the combination of molecular primers and probes is used to quantify the level of gene expression of an inflammatory biomarker in a sample. In one example, the level is expressed as fold change relative to an appropriate control. "fold change" is the ratio of the difference between two quantities. In another example, gene expression levels of inflammatory biomarkers can be measured in a sample using a multiplex PCR assay, for example, a Luminex assay (see, e.g., cook et al methods 158:27-32.2019). In these examples, gene expression of multiple inflammatory biomarkers is measured in a single sample. In another example, gene expression of multiple inflammatory biomarkers is measured in separate samples.
In one example, the level of an inflammatory biomarker is measured in serum. In another example, the level of an inflammatory biomarker is measured in plasma.
In one example, multiple samples are obtained from a subject over time. Inflammatory biomarker levels may be determined in these samples to monitor the level of inflammation in the subject over time. In one example, samples are taken at baseline (i.e., prior to administration of the cell therapy) and after administration of the cell therapy. The levels of the inflammatory biomarkers can be compared between samples to determine whether the levels of the inflammatory biomarkers have been altered (e.g., reduced). In another example, the level of an inflammatory biomarker can be determined in a plurality of samples taken over time (e.g., baseline, day 7, day 14, day 21, day 30). In these examples, the sample may be evaluated to determine whether the inflammation has changed (e.g., abated), and in the case of abated, whether the abation of the inflammation is persistent. In one example, the persistent reduction in inflammation is determined based on a reduction from baseline in inflammation observed in at least two samples after administration of cell therapy. In one example, samples were taken on days 7 and 14. In another example, samples were taken on days 14 and 21. In another example, samples were taken on day 21 and day 30. In another example, samples were taken on day 7, day 14, day 21, and day 30. In one example, a persistent change in inflammation represents a change observed on days 7 and 14 after administration of a cell therapy. In another example, a persistent change in inflammation represents a change observed on days 7, 14, and 21 after administration of a cell therapy. In one example, a persistent change in inflammation is indicative of the changes observed on days 14 and 21 following administration of a cell therapy.
In one example, the level is expressed as fold change relative to an appropriate control. "fold change" is the ratio of the difference between two quantities. In one example, the fold change is calculated as log2 (fold change). In the examples, levels are expressed as fold changes relative to ARDS patients < 65 years old. In another example, the level is expressed as a fold change from a baseline level. In another example, the level is expressed as fold change relative to a patient not suffering from ARDS. In the examples, levels are expressed as fold change relative to ARDS patients < 65 years old and not taking corticosteroids.
In another example, the level of expression is in terms of the amount of a particular marker expressed by a cell of the disclosure under culture conditions. In one example, expression represents cell surface expression. In another example, the level is expressed in terms of how much of a particular marker is released from the cells described herein under culture conditions. In one example, the level is expressed in pg/ml. In another example, the level is in pg/10 6 Individual cells are indicated. If desired, the level of pg/ml may be converted to pg/10 6 Individual cells. For example, in the context of TNF-R1, in one example, 200pg/MlTNF-R1 corresponds to about 23.5pg TNF-R1/10 6 Individual cells. In one example, in the context of TNF-R1, 225pg/ml TNF-R1 corresponds to about 26.5pg TNF-R1/10 in one example 6 Individual cells. In one ofIn an example, 230pg/Ml TNF-R1 corresponds to about 27pg TNF-R1/10 6 Individual cells. In another example, 260pg/ml TNF-R1 corresponds to about 30pg TNF-R1/10 6 Individual cells. In another example, 270pg/ml TNF-R1 corresponds to about 32pg TNF-R1/10 6 Individual cells, etc.
In one example, the level of a particular marker is determined under culture conditions. The term "culture conditions" is used to refer to cells grown in culture. In one example, culture conditions refer to actively dividing cell populations. In one example, such cells may be in an exponential growth phase. For example, the level of a particular marker may be determined by taking a sample of cell culture medium and measuring the level of the marker in the sample. In another example, the level of a particular marker may be determined by taking a sample of cells and measuring the level of the marker in the cell lysate. One skilled in the art will appreciate that secreted markers can be measured by sampling the culture medium, while markers expressed on the cell surface can be measured by evaluating a sample of cell lysate. In one example, the sample is obtained while the cells are in an exponential growth phase. In one example, the sample is obtained after at least two days of culture.
Culturing expanded cells from cryopreserved intermediates means thawing the cells, subjecting them to low temperature freezing, and culturing them in vitro under conditions suitable for cell growth.
In one example, the "level" or "amount" of a particular marker (e.g., TNF-R1) is determined after cells are cryopreserved and then inoculated back into culture. For example, the level is determined after the first cryopreservation of the cells. In another example, the level is determined after the second cryopreservation of the cells. For example, cells may be expanded from a cryopreserved intermediate culture and a second cryopreservation performed prior to re-seeding into the culture so that the level of a particular marker may be determined under culture conditions.
"biomarker" refers to a naturally occurring molecule, gene, or feature that can recognize a particular pathological or physiological process, disease, or the like. As used herein, "inflammatory biomarker" refers to a naturally occurring molecule that indicates the presence or absence of an active inflammatory disease and/or an active inflammatory pathway. In one example, the inflammatory biomarker is a cytokine. In another example, the inflammatory biomarker is a chemokine. Examples of inflammatory biomarkers are discussed further below, and include, for example, CXCL10, CXCL9, CCL2, CCL3, CCL7, IL-6, IL-8, CCL19, IL-2, ferritin, and/or CRP.
By "sustained CRP level" is meant that the subject has CRP levels that are not significantly reduced (e.g., p < 0.05) after treatment with the MLPSC. In one example, sustained CRP levels indicate that the CRP levels of the subject remain (i.e., are not significantly altered) after receiving the first dose of MLPSC.
In one example, the methods of the present disclosure relate to treating patients with elevated inflammatory biomarker levels and/or patients selected for treatment. In one example, the level of the inflammatory biomarker is increased relative to a control population. In one example, the patient's inflammatory biomarker levels are elevated relative to a patient < 65 years old. In another example, the patient's inflammatory biomarker level is elevated relative to its baseline level.
"isolated" or "purified" means a cell that has been separated from at least some components of its natural environment. The term includes the general physical separation of cells from their natural environment (e.g., removal from a donor). The term "isolated" includes altering the relationship of a cell to its directly adjacent cell by, for example, dissociation. The term "isolated" does not refer to cells in a tissue section. When used in reference to a population of cells, the term "isolated" includes a population of cells resulting from proliferation of isolated cells of the present disclosure.
The term "passaging", or "subculture" is used in the context of the present disclosure to refer to known cell culture techniques for maintaining cell survival and growth under culture conditions for a longer period of time such that the number of cells is increasing. The extent of subculturing a cell line is generally expressed as "number of passages" and is generally used to refer to the number of times a cell has been subcultured. In one example, one passage includes removing non-adherent cells and leaving adherent mesenchymal lineage precursors or stem cells. Such mesenchymal lineage precursor or stem cells can then be dissociated from the matrix or flask (e.g., by using a protease, such as trypsin or collagenase), a medium can be added, optional washing can be performed (e.g., by centrifugation), and then the mesenchymal lineage precursor or stem cells can be re-plated or re-inoculated into one or more culture vessels having a greater total surface area. The mesenchymal lineage precursor or stem cells can then continue to expand in culture. In another example, the method of removing non-adherent cells includes a step of non-enzymatic treatment (e.g., with EDTA). In one example, the mesenchymal lineage precursor or stem cells are passaged at or near confluence (e.g., about 75% to about 95% confluence). In one example, mesenchymal lineage precursor or stem cells are seeded at a concentration of about 10%, about 15%, or about 20% cells/m 1 medium.
The term "medium" or "medium" as used in the context of the present disclosure includes components of the environment surrounding the cells in culture. It is contemplated that the medium facilitates and/or provides conditions suitable for cell growth. The medium may be a solid, a liquid, a gas or a mixture of phases and materials. The medium may include a liquid growth medium and a liquid medium that does not sustain cell growth. Exemplary gaseous media include the gas phase to which cells grown on a petri dish or other solid or semi-solid support are exposed.
As used herein, the term "treating" includes administration of a population of mesenchymal lineage stem cells or precursor cells and/or progeny thereof and/or soluble factors derived therefrom, thereby alleviating or eliminating at least one ARDS symptom. In one example, the treatment comprises administering a population of culture expanded mesenchymal stem cells or precursor cells. In one example, the treatment response is determined relative to a baseline. In one example, the therapeutic response is determined relative to a control patient population. In one example, the treatment improves ARDS in the subject from severe to moderate. In one example, the treatment improves respiratory function. In one example, the treatment allows the subject to be free of mechanical ventilation.
In one example, the methods of the present disclosure inhibit disease progression or disease complications in a subject. By "inhibiting" disease progression or disease complications in a subject is meant preventing or reducing disease progression and/or disease complications in the subject. Thus, in one example, the methods of the present disclosure inhibit progression of ARDS severity.
As used herein, the term "preventing" includes administration of a population of mesenchymal stem cells or precursor cells and/or their progeny and/or soluble factors derived therefrom, thereby preventing or inhibiting the development of at least one symptom of ARDS.
As used herein, the term "subject" refers to a human subject. For example, the subject may be an adult. Terms such as "subject," "patient," or "individual" are terms that may be used interchangeably in this disclosure in context.
The term "thrombosis" as used herein refers to the formation of a thrombus or blood clot. In one example, thrombosis is "arterial thrombosis" in which a blood clot forms in an artery. Such blood clots are particularly dangerous to the subject because they can obstruct blood flow to major organs such as the heart or brain. In one example, thrombosis is "venous thrombosis" in which a blood clot forms in a vein.
The term "pulmonary embolism" as used herein refers to the blockage of a pulmonary artery by substances that migrate from other parts of the body through the blood.
As used herein, the term "genetically unmodified" refers to cells that have not been modified by nucleic acid transfection. For the avoidance of doubt, in the context of the present disclosure, mesenchymal lineage precursors or stem cells transfected with a nucleic acid encoding Ang1 will be considered genetically modified.
The term "total dose" is used in the context of the present disclosure to refer to the total number of cells received by a subject treated according to the present disclosure. In one example, the total dose consists of one cell administration. In another example, the total dose consists of two cell administrations. In another example, the total dose consists of three cell administrations. In another example, the total dose consists of four or more cell administrations. For example, the total dose may consist of two to five cellular administrations. In each example, the total dose exceeded 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg). For example, a total dose of over 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg) can be administered by three doses. In this example, the subject may have received 200 ten thousand prior doses of mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg).
The term "clinically proven" (independently used or modified term "effective") means that efficacy has been demonstrated by clinical trials that have met the approval criteria of the U.S. food and drug administration, EMEA, or corresponding national regulatory authorities. For example, the clinical study may be a randomized, double-blind study of sufficient scale for clinical demonstration of the efficacy of the composition. In one example, a clinically proven effective amount is an amount that the clinical trial shows to meet the specified endpoint. In one example, the endpoint is protection from death.
Thus, the terms "clinically proven efficacy" and "clinically proven effective" may be used in the context of the present disclosure to refer to dosages, dosage regimens, treatments or methods disclosed herein. Efficacy may be measured based on changes in the course of a disease in response to administration of the compositions disclosed herein. For example, the compositions of the present disclosure are administered to a subject in an amount and for a duration sufficient to cause an improvement, preferably a persistence improvement, in at least one indicator reflecting the severity of ARDS. Various indicators reflecting the severity of ARDS can be evaluated to determine if the amount and time of treatment is sufficient. Such indicators include, for example, clinically recognized indicators of disease severity or symptoms. In one example, the degree of improvement is determined by a physician, who may make this determination based on signs, symptoms, or other test results. In one example, clinically effective amounts have been shown to increase patient survival. In another example, a clinically proven effective amount reduces the risk of mortality in a subject. In another example, a clinically proven effective amount reduces circulating CRP levels in a subject. In another example, a clinically proven effective amount improves respiratory function in a subject.
Throughout this specification, the word "comprise" or variations such as "comprises" or "comprising" will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps.
As used herein, the singular forms "a," "an," and "the" include the singular and plural references unless the context clearly dictates otherwise.
Acute Respiratory Distress Syndrome (ARDS)
The methods of the present disclosure relate to the treatment of Acute Respiratory Distress Syndrome (ARDS) by administration of the compositions disclosed herein. In one example, the method comprises administering a composition comprising an MLPSC. Thus, in one example, the composition may comprise an MSC.
The term "Acute Respiratory Distress Syndrome (ARDS)" is a type of respiratory failure characterized by extensive inflammation, poor oxygenation and inflexibility or "stiff" lungs. The condition is typically associated with capillary endothelial injury and diffuse alveolar injury.
In one example, the methods of the present disclosure prevent or treat a subject with mild ARDS. In another example, the methods of the present disclosure prevent or treat a subject with moderate ARDS. In another example, the methods of the present disclosure prevent or treat a subject with severe ARDS. In another example, the methods of the present disclosure prevent or treat a subject with moderate or severe ARDS. In one example, the methods of the present disclosure treat a subject with ARDS in need of ventilation. For example, the subject may rely on a mechanical ventilator.
In one example, the severity of ARDS is diagnosed based on the PaO2/FiO2 ratio. For example, the severity of ARDS can be diagnosed as follows: (light: 26.6kPa < PaO2/FiO 2.ltoreq.39.9 kPa; medium: 13.3kPa < PaO2/FiO 2.ltoreq.26.6 kPa; heavy: paO2/FiO 2.ltoreq.13.3 kPa). In one example, the severity of ARDS can be diagnosed according to the berlin standard, as summarized in the following table:
/>
in another example, the severity of ARDS may be diagnosed as follows: mild (PaO 2/FiO2200 to 300 mmHg); moderate (PaO 2/FiO2100 to 200 mmHg); severe (PaO 2/FiO2, less than 100 mmHg).
In one example, a subject treated according to the methods of the present disclosure is greater than or equal to 65 years old.
In another example, a subject treated according to the methods of the present disclosure is taking a corticosteroid. In one example, the corticosteroid is a long-acting or medium-acting (half-life < 36 hours) corticosteroid. In one example, the corticosteroid is long-acting (half-life of 36 to 72 hours). In one example, the corticosteroid is dexamethasone. Other examples of corticosteroids include prednisone and methylprednisolone. For example, the subject may take dexamethasone. In one example, the subject is greater than or equal to 65 years old and is taking a corticosteroid such as dexamethasone.
In one example, the methods of the present disclosure include administering a cell composition disclosed herein, such as a composition comprising an MLPSC. In another example, the methods of the present disclosure comprise administering a cell composition disclosed herein and a corticosteroid. In this example, the corticosteroid may be administered simultaneously or sequentially with the cell composition. In one example, the subject has previously taken a corticosteroid prior to administration of the cell compositions disclosed herein. In this example, the corticosteroid may be continued to be administered along with the cell composition.
A subject treated according to the present disclosure may have symptoms that are indicative of ARDS. Exemplary symptoms may include fatigue, dyspnea, shortness of breath, inability to exercise or reduced exercise capacity, coughing with or without blood or mucus, pain during inspiration or expiration, wheezing, chest distress, weight loss of unknown origin, musculoskeletal pain, accelerated breathing (shortness of breath), and blue skin (cyanosis).
In another example, the subject has pneumonia.
In another example, the subject has ARDS secondary to a viral infection. In one example, the ARDS of the subject is secondary to infection by a rhinovirus, influenza virus, respiratory Syncytial Virus (RSV) or coronavirus. In one example, the ARDS of the subject is secondary to an infection with a coronavirus. For example, ARDS of a subject may be secondary to infection with SARS-CoV, MERS-CoV or COVID-19. In one example, the ARDS of the subject is secondary to SARS-CoV-2 infection.
In one example, the subject has one or more of myocarditis, pericarditis, or valvulitis. In one example, the subject has viral myocarditis, pericarditis, or valvulitis. For example, the subject may have viral myocarditis.
In one example, ARDS is caused by a viral infection. For example, ARDS can be caused by rhinoviruses, influenza viruses, respiratory Syncytial Viruses (RSV) or coronaviruses. In one example, ARDS may be caused by coronaviruses. For example, the coronavirus may be a coronavirus (SARS-CoV), a middle east respiratory syndrome coronavirus (MERS-CoV), or a COVID-19. In one example, the ARDS is caused by Ai Bashi (Epstein-Barr) virus (EBV) or Herpes Simplex Virus (HSV).
In another example, ARDS is caused by thrombosis. In another example, ARDS is caused by embolism. In one example, ARDS is caused by pulmonary embolism.
In another example, ARDS is secondary to hemophagocytic lymphoproliferative disorder (HLH). HLH is a life threatening disease characterized by excessive inflammation of lymphocytes and macrophages. HLH may be triggered by a viral infection (such as EBV, CMV, HHV). Thus, in one example, HLH is secondary or acquired HLH. For example, HLH can be secondary to a viral infection and lead to the development of ARDS in a subject.
In one example, the treatment is protected from death or imparts improved survival. In one example, protection from death is determined 60 days after treatment. In another example, protection from death is determined 50 to 70 days after treatment. For example, the risk of mortality in a treated subject may be reduced after treatment. In one example, the reduction in mortality risk in the treated subject is between 30% and 60%. In one example, the reduction in mortality risk in the treated subject is between 40% and 50%. In one example, the risk of mortality in the treated subject is reduced by at least 30%. In one example, the risk of mortality in the treated subject is reduced by at least 40%.
In one example, the treatment improves oxygenation of the subject. In one example, the improved oxygenation corresponds to a change in the berlin standard. In one example, the treatment improves oxygenation to a level corresponding to a more mild grade of ARDS. In one example, the treatment improves oxygenation levels from severe to moderate.
In one example, treatment according to the methods of the present disclosure reduces the risk of thrombosis in a subject. In one example, the risk of the subject is reduced relative to a subject not receiving the treatment. In one example, the treatment reduces the risk that thrombosis is arterial thrombosis. Thus, in one example, the treatment reduces the risk of heart attack or stroke in a subject with ARDS.
In another example, the disclosure encompasses selecting a subject with ARDS for treatment. In one example, the subject has moderate or severe ARDS. In one example, the method comprises selecting a subject with ARDS greater than or equal to 65 years old. In one example, the method comprises selecting a subject greater than or equal to 65 years old who is taking a corticosteroid. In one example, a selected subject is treated according to the methods disclosed herein.
Inflammatory organismsMarker(s)
The methods of the present disclosure relate to selecting and treating patients with ARDS with elevated levels of inflammatory biomarkers. Inflammatory biomarkers are associated with upregulation of inflammatory pathways, which are associated with increased disease severity.
In one example, the inflammatory biomarker is indicative of increased neutrophil and macrophage influx into the lung, e.g., CCR2 binding chemokines. The C-C chemokine receptor type 2 (CCR 2) is a G protein coupled receptor, the ligands of which include the Monocyte Chemotactic Protein (MCP) family of chemokines, including CCL2 (C-C motif chemokine ligand 2), CCL3 (C-C motif chemokine ligand 3) and CCL7 (C-C motif chemokine ligand 7). Elevated levels of CCR 2-binding chemokines are thought to result in greater attraction of CXCR3 neutrophils and neutrophil precursors to the lung. Thus, in one example, the CCL2 level of the subject is elevated. In another example, the subject has elevated CCL3 levels. In another example, the subject has elevated CCL7 levels.
In another example, the inflammatory biomarker that indicates increased neutrophil and macrophage influx into the lung is CXCR3 (C-X-C motif chemokine receptor 3) binding chemokine. Examples of CXCR 3-binding chemokines include C-X-C motif chemokine ligand 10 (CXCL 10) and C-X-C motif chemokine ligand 9 (CXCL 9). In another example, the subject has elevated CXCL10 levels. In another example, the subject has elevated CXCL9 levels.
In another example, an inflammatory biomarker is indicative of increased macrophage inflammation and increased neutrophil migration to the lung, e.g., interleukin-6 (IL-6) and interleukin-8 (IL-8). In one example, the IL-6 level in the subject is increased. In one example, the subject has an elevated IL-8 level.
In another example, an inflammatory biomarker is indicative of T cell activation/proliferation and apoptotic death, e.g., C-C motif chemokine ligand (CCL 19) and interleukin-2 (IL-2). In one example, the subject has elevated CCL19 levels. In one example, the subject has an elevated IL-2 level.
The level of inflammatory biomarkers can also be reduced after treatment with the compositions of the present disclosure. In one example, a decrease in inflammatory biomarkers is indicative of decreased neutrophil and macrophage influx into the lung, e.g., CCR2 binding chemokines such as CCL2, CCL3, and CCL7. In one example, the subject's CCL2 level is reduced. In another example, the subject's CCL3 level is decreased. In another example, the subject's CCL7 level is decreased. In another example, the inflammatory biomarker is a CXCR 3-binding chemokine, such as CXCL10 and CXCL9. In one example, the subject has a reduced CXCL10 level. In another example, the subject's CXCL9 level is reduced.
In another example, a decrease in an inflammatory biomarker is indicative of decreased inflammatory body. The inflammasome is a stimulus-induced cytoplasmic polyprotein complex. In another example, a decrease in an inflammatory biomarker is indicative of decreased macrophage activation and neutrophil homing to the lung. When reduced, examples of inflammatory biomarkers that indicate inflammatory body reduction and macrophage activation/neutrophil homing to the lung reduction are IL-6, IL-8, tumor Necrosis Factor (TNF), and interleukin-18 (IL-18). In one example, the subject has a reduced IL-6 level. In another example, the subject has a reduced IL-8 level. In another example, the TNF level of the subject is decreased. In another example, the subject has a reduced IL-18 level.
In another example, a decrease in an inflammatory biomarker is indicative of decreased T cell influx and activation. Examples of inflammatory biomarkers indicate reduced T cell influx and activation when the C-C motif chemokine ligand 19 (CCL 19), interleukin-4 (IL-4), interleukin-13 (IL-13) and granulocyte-macrophage colony stimulating factor (GM-CSF) are reduced. In one example, the subject has a reduced level of CCL 19. In another example, the subject has a reduced IL-4 level. In another example, the subject has a reduced IL-13 level. In another example, the subject has a reduced level of GM-CSF.
In another example, a decrease in inflammatory biomarkers is indicative of a decrease in circulating biomarkers of macrophage and neutrophil inflammation, e.g., CRP, ferritin, or D-dimer. In one example, the reduced circulating biomarker is "C-reactive protein" or "CRP. CRP is an inflammatory mediator whose level increases in the event of recurrence of acute inflammation and returns to normal rapidly after resolution of inflammation. Circulating CRP levels can be measured in plasma samples to provide a measure of inflammation in a subject.
In one example, the treatment reduces CRP levels in the subject. In one example, the treatment reduces CRP by at least 100mg/d1 as compared to baseline. In another example, the treatment reduces CRP by at least 150mg/dl as compared to baseline. In one example, the treatment reduces CRP levels in the subject by about 0.1-fold. In another example, the treatment reduces CRP levels in the subject by about 0.2-fold. In another example, the treatment reduces CRP levels in the subject by about 0.3-fold. In another example, the treatment reduces CRP levels in the subject by about 0.4-fold. In another example, the treatment reduces CRP levels in the subject by about 0.5-fold. In another example, the treatment reduces CRP levels in the subject by a factor of 0.6. In these examples, the treatment reduces CRP levels in the subject by a fold change relative to baseline CRP levels in the subject.
In an example, the subject has sustained CRP levels after administration of the first dose of MLPSC. In one example, the subject has sustained CRP levels 7 days after administration of the first dose of MLPSC. In one example, the subject has sustained CRP levels 14 days after administration of the first dose of MLPSC. In one example, the subject has sustained CRP levels 21 days after administration of the first dose of MLPSC. In one example, the subject has sustained CRP levels 30 days after administration of the first dose of MLPSC.
In another example, the reduced circulating biomarker is ferritin. Ferritin is a blood protein containing iron. Ferritin levels may be measured in a blood sample to provide a measure of the subject's iron level. In one example, the treatment reduces ferritin levels in the subject. In one example, the treatment reduces ferritin levels in the subject by a factor of about 0.1. In another example, the treatment reduces ferritin levels in the subject by a factor of about 0.2. In another example, the treatment reduces ferritin levels in the subject by a factor of about 0.3. In another example, the treatment reduces ferritin levels in the subject by about 0.4-fold. In another example, the treatment reduces ferritin levels in the subject by a factor of about 0.5. In these examples, the treatment reduces ferritin levels in the subject by a fold change relative to baseline ferritin levels in the subject.
Methods of determining the levels of inflammatory biomarkers disclosed herein are known in the art. In one example, the level of a particular marker is determined in a sample obtained from a patient or subject (e.g., a blood sample, a plasma sample, or a serum sample). For example, the level of an inflammatory biomarker according to the present disclosure is determined in a plasma sample. In another example, the level of an inflammatory biomarker is determined in a serum sample.
In one example, the level of an inflammatory biomarker is determined by measuring the level of protein expression in a sample obtained from a subject. For example, inflammatory biomarker levels may be measured in a sample using an antibody-based immunoassay (e.g., an enzyme-linked immunosorbent (ELISA)) assay, a multiplex immunoassay (e.g., a Luminex assay) (see, e.g., cook et al methods 158:27-32.2019), or a fluorescent bead-based immunoassay. In these examples, the level of inflammatory biomarker is expressed in pg/mL. In another example, the level of an inflammatory biomarker is expressed as a fold change relative to an appropriate control. In another example, the level of an inflammatory biomarker is determined by measuring the level of gene expression in a sample obtained from a subject. For example, the inflammatory biomarker levels may be measured in a sample using a molecular-based assay (e.g., a qualitative Polymerase Chain Reaction (PCR) based assay) or a multiplex PCR assay (e.g., a Luminex assay) (see, e.g., cook et al methods.158:27-32.2019). In one example, the gene expression level of an inflammatory biomarker is expressed as fold change relative to an appropriate control. For example, the fold change is calculated as log2 (fold change).
In the examples, the level of inflammatory biomarkers is expressed as fold change relative to ARDS patients < 65 years old.
In one example, the level of inflammatory biomarkers is increased by more than 1 fold relative to ARDS patients < 65 years old. In another example, the level of the inflammatory biomarker is increased by more than a factor of 1 relative to the baseline level. In another example, the level of inflammatory biomarker is increased by more than 1 fold relative to a patient not having ARDS. In another example, the level of inflammatory biomarker is increased by at least 2-fold relative to an ARDS patient < 65 years old. In another example, the level of the inflammatory biomarker is increased by at least 2-fold relative to the baseline level. In another example, the level of the inflammatory biomarker is increased by at least 2-fold relative to a patient not having ARDS. In another example, the level of inflammatory biomarker is increased by at least 5-fold relative to an ARDS patient < 65 years old. In another example, the level of the inflammatory biomarker is increased by at least 5-fold relative to the baseline level. In another example, the level of the inflammatory biomarker is increased by at least 5-fold relative to a patient not having ARDS. In another example, the level of inflammatory biomarker is increased by more than 5-fold relative to a patient not having ARDS.
In one example, the levels of multiple inflammatory biomarkers are measured in a single sample. In another example, the levels of multiple inflammatory biomarkers are measured in separate samples. In one example, the level of the biomarker is measured prior to treatment with the MLPSC. In another example, the level of the biomarker is measured after treatment with the MLPSC. In another example, the level of the biomarker is measured at baseline and monitored over time to determine if the subject requires higher doses or longer administrations.
The levels of the inflammatory biomarkers between samples can be compared to determine if the levels of the inflammatory biomarkers are reduced. In these examples, the sample may be evaluated to determine whether the inflammation has been reduced and whether the reduction in inflammation is persistent. In one example, the persistent reduction in inflammation is determined based on a reduction from baseline in inflammation observed in at least two samples after administration of cell therapy.
Methods of treating ARDS patient populations
In one example, the methods of the present disclosure relate to treating an ARDS patient population. In another example, the methods of the present disclosure relate to selecting an ARDS patient for treatment according to the methods disclosed herein.
In one example, the method comprises selecting a subject with an increased inflammatory biomarker indicative of: increased flow of neutrophils and macrophages into the lung; macrophage inflammation and neutrophil migration to the lung are increased; and/or T cell activation/proliferation and apoptotic death, and administering to the subject a composition comprising more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg).
In another example, the method comprises selecting a subject with ARDS greater than or equal to 65 years old and administering to the subject a composition comprising more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg).
In another example, the method comprises administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg) compositions, wherein the subject is greater than or equal to 65 years old and/or has an increased inflammatory biomarker that indicates increased neutrophil and macrophage influx into the lung; macrophage inflammation and neutrophil migration to the lung are increased; and/or T cell activation/proliferation and apoptotic death.
In another example, the method includes:
-determining or having determined the level of one or more inflammatory biomarkers of the subject, said inflammatory biomarkers being selected from the list comprising: (i) CXCR3 binds to chemokines, preferably CXCL10 and/or CXCL9; (ii) CCR 2-binding chemokines, preferably CCL2, CCL3 and/or CCL7; (iii) IL-6; (v) IL-8; (vi) CCL19; (vii) IL-2; and/or (viii) CRP;
-selecting a subject greater than or equal to 65 years old and/or having elevated levels of one or more inflammatory biomarkers and/or sustained CRP levels 7 days after administration of the first dose of mesenchymal lineage precursor or stem cells (MLPSC); and
-administering to the subject a composition comprising more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg).
In another example, a cell therapy is administered to a subject and monitored to determine whether an inflammatory biomarker is reduced in response to the treatment. If the inflammatory biomarker is free from reduction, another dose of cells is administered to the subject. In one example, 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg) are administered to a subject with ARDS, and monitoring is performed to determine if inflammatory biomarkers are reduced. In one example, the subject is monitored by determining the subject's inflammatory biomarker level prior to administration of the MLPSC, and then determining the subject's inflammatory biomarker level after administration of the MLPSC. In one example, the inflammatory biomarker is selected from the group consisting of: CXCL10, CXCL9, CCL2, CCL3, CCL7, IL-6, IL-8, CCL19, IL-2, ferritin and/or CRP. In one example, the biomarker is CRP. In another example, the biomarker is ferritin. In one example, the level of the biomarker is not reduced relative to the baseline level at day 7 post-treatment. In another example, the level of the biomarker is not reduced relative to the baseline level on day 14 post-treatment. In one example, if the level of one or more biomarkers does not decrease from baseline, another dose of MLPSC is administered to the subject. In one example, another dose comprises 200 ten thousand MLPSCs per kilogram body weight (cells/kg). In one example, another dose is administered on day 7 after the initial treatment. In another example, another dose is administered on day 14 after the initial treatment. In another example, another dose is administered on day 21 after the initial treatment. In another example, two or more additional doses are administered. For example, the subject may receive another dose on day 7, day 14, and day 21 after the initial treatment. In another example, another dose is administered until the level of inflammatory biomarker is reduced relative to baseline. In another example, a subject may receive two doses of 300 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg). For example, doses may be administered on days 7 and 14.
Accordingly, in one example, the present disclosure relates to a method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising:
-administering 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg) to the subject;
-determining or having determined the subject level of one or more biomarkers selected from the group consisting of: CXCL10, CXCL9, CCL2, CCL3, CCL7, IL-6, IL-8, CCL19, IL-2, ferritin and/or CRP;
if the level of one or more biomarkers does not decrease from baseline, then an additional dose of MLPSC is administered to the subject, wherein after the additional dose, more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg) have been administered to the subject.
In one example, the treatment improves respiratory function in the subject. In one example, improved respiratory function is defined as regression and/or improvement of ARDS as defined by berlin standards. In one example, improved blood oxygenation indicates improved respiratory function. In an example, the treatment improves respiratory function on day 14. In another example, the treatment improves respiratory function on day 21.
Mesenchymal precursor cells
As used herein, the term "mesenchymal lineage precursor or stem cells (MLPSC)" refers to undifferentiated pluripotent cells that have the ability to self-renew while retaining multipotency and to differentiate into many cell types of mesenchymal origin (e.g., osteoblasts, chondrocytes, adipocytes, stromal cells, fibroblasts, and tendons) or non-mesoderm origin (e.g., hepatocytes, neural cells, and epithelial cells). For the avoidance of doubt, "mesenchymal lineage precursor cells" refers to cells capable of differentiating into mesenchymal cells, such as bone, cartilage, muscle and adipocytes, and fibrous connective tissue.
The term "mesenchymal lineage precursor or stem cells" includes the parent cell and its undifferentiated progeny. The term also includes mesenchymal precursor cells, pluripotent stromal cells, mesenchymal Stem Cells (MSCs), perivascular mesenchymal precursor cells and their undifferentiated progeny.
The mesenchymal lineage precursor or stem cells can be autologous, allogeneic, xenogeneic, syngeneic, or isogenic. Autologous cells are isolated from the same body to be re-implanted. Allogeneic cells are isolated from a donor of the same species. The xenogeneic cells are isolated from a donor of another species. Isogenic or isogenic cells are isolated from genetically identical organisms, such as twins, clones or highly inbred research animal models.
In one example, the mesenchymal lineage precursor or stem cells are allogeneic. In one example, allogeneic mesenchymal lineage precursor or stem cells are culture expanded and cryopreserved.
Mesenchymal lineage precursor or stem cells are found predominantly in bone marrow, but are also shown to be present in a variety of host tissues including, for example, umbilical cord blood and cord, adult peripheral blood, adipose tissue, trabecular bone, and dental pulp. They are also found in skin, spleen, pancreas, brain, kidney, liver, heart, retina, brain, hair follicle, intestine, lung, lymph node, thymus, ligament, tendon, skeletal muscle, dermis and periosteum; and is capable of differentiating into a germ line, such as mesoderm and/or endoderm and/or ectoderm. Thus, mesenchymal lineage precursor or stem cells can differentiate into a number of cell types including, but not limited to, fat, bone, cartilage, elastic cartilage, muscle, and fibrous connective tissue. The particular lineage commitment and differentiation pathway that these cells enter depends on various effects from mechanical influences and/or endogenous bioactive factors (e.g., growth factors, cytokines) and/or local microenvironmental conditions established by the host tissue.
The terms "enriched", "enriched" or variants thereof are used herein to describe a population of cells in which the proportion of one particular cell type or the proportion of a plurality of particular cell types is increased when compared to an untreated population of cells (e.g., cells in a natural environment). In one example, the population enriched for mesenchymal lineage precursor or stem cells comprises at least about 0.1% or 0.5% or 1% or 2% or 5% or 10% or 15% or 20% or 25% or 30% or 50% or 75% mesenchymal lineage precursor or stem cells. In this regard, the term "cell population enriched for mesenchymal lineage precursor or stem cells" will be considered to provide explicit support for the term "cell population comprising X% mesenchymal lineage precursor or stem cells", where X% is a percentage as described herein. In some examples, the mesenchymal lineage precursor or stem cells can form clonogenic colonies, e.g., CFU-F (fibroblasts), or a subset thereof (e.g., 50% or 60% or 70% or 90% or 95%) can have such activity.
In one example of the present disclosure, the mesenchymal lineage precursor or stem cells are Mesenchymal Stem Cells (MSCs). MSCs may be of homogeneous composition or may be a mixed population of cells enriched in MSCs. Homogeneous MSC compositions can be obtained by culturing adherent bone marrow or periosteal cells, and MSCs can be identified by specific cell surface markers identified with unique monoclonal antibodies. Methods for obtaining a population of cells enriched in MSCs are described, for example, in U.S. Pat. No. 5,486,359. Alternative sources of MSCs include, but are not limited to, blood, skin, cord blood, muscle, fat, bone, and perichondrium. In one example, the MSC is allogeneic. In one example, MSCs are cryopreserved. In one example, MSCs are culture expanded and cryopreserved.
In another example, the mesenchymal lineage precursor or stem cell is cd29+, cd54+, cd73+, cd90+, cd102+, cd105+, cd106+, cd166+, MHC1+ MSC.
In one example, mesenchymal lineage precursor or stem cells are culture expanded from a population of MSCs expressing markers (including CD73, CD90, CD105, and CD 166) and lacking expression of hematopoietic cell surface antigens (e.g., CD45 and CD 31). For example, mesenchymal lineage precursor or stem cells can be culture expanded from a population of MSCs (CD73+, CD90+, CD105+, CD166+, CD 45-and CD 31-). In one example, the population of MSCs is further characterized by low levels of Major Histocompatibility Complex (MHC) class I. In another example, MSCs are negative for major histocompatibility complex class II molecules and negative for co-stimulatory molecules CD40, CD80 and CD 86. In one example, the culture amplification includes 5 generations.
In one example, the mesenchymal lineage precursor or stem cells are cd105+, cd156+, and CD45-. In another example, the mesenchymal lineage or precursor cell also expresses TNFR1 and inhibits IL-2Rα expression on activated lymphocytes.
Isolated or enriched mesenchymal lineage precursor or stem cells can be expanded in vitro by culture. Isolated or enriched mesenchymal lineage precursor or stem cells can be cryopreserved, thawed, and subsequently expanded in vitro by culture.
In one example, isolated or enriched mesenchymal lineage precursor or stem cells are seeded at 50,000 viable cells/cm 2 in a medium (serum-free or serum-supplemented), e.g., alpha minimal medium (alpha MEM) supplemented with 5% Fetal Bovine Serum (FBS) and glutamine, and allowed to warm at 37 ℃, 20% O 2 Adhere to the culture vessel overnight. The medium is then replaced and/or changed as required and the cells are incubated at 37℃with 5% O 2 The culture was continued for 68 to 72 hours.
As will be appreciated by those skilled in the art, cultured mesenchymal lineage precursor or stem cells are phenotypically different from in vivo cells. For example, in one embodiment, they express one or more of the following markers: CD44, NG2, DC146, and CD140b. The cultured mesenchymal lineage precursor or stem cells are also biologically different from in vivo cells, and have higher proliferation rates than most non-circulating (resting) cells in vivo.
In one example, the population of cells is enriched from a cell preparation comprising STRO-1+ cells in a selectable form. In this regard, the term "selectable form" is understood to mean that the cells express a marker (e.g., a cell surface marker) that allows selection of STRO-1+ cells. The marker may be STRO-1, but is not necessarily. For example, as described herein and/or exemplified herein, cells expressing STRO-2 and/or STRO-3 (TNAP) and/or STRO-4 and/or VCAM-1 and/or CD146 and/or 3G5 (e.g., mesenchymal precursor cells) also express STRO-1 (and may be STRO-1 bright). Thus, the indication that the cell is STRO-1+ does not mean that the cell is selected by STRO-1 expression alone. In one example, cells are selected based on at least STRO-3 expression, e.g., they are STRO-3+ (TNAP+).
References to selection of cells or populations thereof do not necessarily require selection from a particular tissue source. STRO-1+ cells may be selected from or isolated or enriched from a variety of sources, as described herein. That is, in some examples, these terms provide support for selection from any tissue or vascularized tissue comprising STRO-1+ cells (e.g., mesenchymal precursor cells) or tissue comprising pericytes (e.g., STRO-1+ pericytes) or any one or more of the tissues listed herein.
In one example, the cells used in the present disclosure express one or more markers selected individually or collectively from the group consisting of: TNAP+, VCAM-1+, THY-1+, STRO-2+, STRO-4+ (HSP-90 beta), CD45+, CD146+, 3G5+, or any combination thereof.
"singly" means that the present disclosure individually encompasses the listed markers or sets of markers, and that although individual markers or sets of markers may not be individually listed herein, the appended claims may define such markers or sets of markers individually and dividedly from each other.
"collectively" means that the present disclosure encompasses any number or combination of enumerated markers or sets of markers, and that although the number or combination of such markers or sets of markers may not be specifically listed herein, the appended claims may define such combinations or sub-combinations separately or separately from any other combination of markers or sets of markers.
As used herein, the term "TNAP" is intended to encompass all isoforms of tissue-nonspecific alkaline phosphatase. For example, the term encompasses liver isotype (LAP), bone isotype (BAP) and kidney isotype (KAP). In one example, the TNAP is BAP. In one example, TNAP as used herein refers to a molecule that can bind STRO-3 antibodies produced by a hybridoma cell line deposited with ATCC under the provisions of the budapest treaty at 12 months 19 of 2005 under deposit accession number PTA-7282.
Furthermore, in one example, STRO-1+ cells are capable of producing clonogenic CFU-F.
In one example, a significant proportion of STRO-1+ cells are capable of differentiating into at least two different lineages. Non-limiting examples of lineages in which STRO-1+ cells can committed include bone precursor cells; hepatocyte progenitor cells, multipotent for bile duct epithelial cells and hepatocytes; neural restricted cells, which can produce glial cell precursors and develop into oligodendrocytes and astrocytes; a neuronal precursor that develops into a neuron; myocardium and precursors of cardiomyocytes, glucose-responsive insulin secreting pancreatic beta cell lines. Other lineages include, but are not limited to, odontoblasts and chondrocytes, as well as precursor cells of: retinal pigment epithelial cells, fibroblasts, skin cells such as keratinocytes, dendritic cells, hair follicle cells, renal tubular epithelial cells, smooth and skeletal muscle cells, testicular progenitor cells, vascular endothelial cells, tendons, ligaments, cartilage, adipocytes, fibroblasts, bone marrow stroma, cardiac muscle, smooth muscle, skeletal muscle, pericytes, blood vessels, epithelium, glia, neurons, astrocytes and oligodendrocytes.
In one example, mesenchymal lineage precursor or stem cells are obtained from a single donor, or multiple donors, where these donor samples or mesenchymal lineage precursor or stem cells are then pooled and then culture expanded.
Mesenchymal lineage precursor or stem cells encompassed by the present disclosure can also be cryopreserved prior to administration to a subject. In one example, mesenchymal lineage precursor or stem cells are culture expanded and cryopreserved prior to administration to a subject.
In one example, the present disclosure encompasses mesenchymal lineage precursor or stem cells and their progeny, soluble factors derived therefrom, and/or extracellular vesicles isolated therefrom. In another example, the disclosure encompasses mesenchymal lineage precursor or stem cells and extracellular vesicles isolated therefrom. For example, mesenchymal precursor lineages or stem cells of the present disclosure may be culture expanded for a period of time and under conditions suitable for secretion of extracellular vesicles into the cell culture medium. The secreted extracellular vesicles can then be obtained from the culture medium for use in therapy.
As used herein, the term "extracellular vesicles" refers to lipid particles that are naturally released from cells and range in size from about 30nm to as large as 10 microns, but typically their size is less than 200nm. They may comprise cells derived from release (e.g., mesenchymal stem cells; STRO-1) + Cells), proteins, nucleic acids, lipids, metabolites or organelles.
As used herein, the term "exosomes" refers to a class of extracellular vesicles that are typically in the range of about 30nm to about 150nm in size, and originate from the endosomal compartments of mammalian cells from which they are transported to the cell membrane and released. They may contain nucleic acids (e.g., RNA; microRNAs), proteins, lipids, and metabolites, and play a role in intercellular communication by being secreted from one cell and taken up by other cells to deliver their substances.
Culture expansion of cells
In one example, mesenchymal lineage precursor or stem cells are expanded in culture. The "culture expanded" mesenchymal lineage precursor or stem cell medium differs from freshly isolated cells in that they have been cultured and passaged (i.e., subcultured) in cell culture media. In one example, the culture-expanded mesenchymal lineage precursor or stem cells are culture-expanded for about 4-10 passages. In one example, the mesenchymal lineage precursor or stem cells are expanded in culture for at least 5, at least 6, at least 7, at least 8, at least 9, at least 10 passages. For example, mesenchymal lineage precursor or stem cells can be expanded in culture for at least 5 passages. In one example, the mesenchymal lineage precursor or stem cells can be expanded in culture for at least 5-10 passages. In one example, the mesenchymal lineage precursor or stem cells can be expanded in culture for at least 5-8 passages. In one example, the mesenchymal lineage precursor or stem cells can be expanded in culture for at least 5-7 passages. In one example, mesenchymal lineage precursor or stem cells can be expanded in culture for more than 10 passages. In another example, mesenchymal lineage precursor or stem cells can be expanded in culture for more than 7 passages. In these examples, stem cells can be expanded in culture prior to cryopreservation to provide an intermediate cryopreserved MLPSC population. In one example, the compositions of the present disclosure are prepared from an intermediate cryopreserved MLPSC population. For example, as discussed further below, the expanded intermediate cryopreserved MLPSC population can be further cultured prior to administration. Thus, in one example, mesenchymal lineage precursor or stem cells are culture expanded and cryopreserved. In embodiments of these examples, the mesenchymal lineage precursor or stem cells can be obtained from a single donor or multiple donors, where these donor samples or mesenchymal lineage precursors or stem cells are then pooled and then culture expanded. In one example, the culture amplification process comprises:
Expanding the number of living cells by passaging to provide a preparation of at least about 10 hundred million living cells, wherein the passaging comprises establishing a primary culture of isolated mesenchymal lineage precursor or stem cells, and then continuously establishing a first non-primary (P1) culture of mesenchymal lineage precursor or stem cells isolated from a previous culture;
expanding the isolated P1 culture of mesenchymal lineage precursor or stem cells into a second non-primary (P2) culture of mesenchymal lineage precursor or stem cells by passaging expansion; and, in addition, the processing unit,
preparing and cryopreserving an intermediate mesenchymal lineage precursor or stem cell preparation in the process obtained from a P2 culture of mesenchymal lineage precursor or stem cells; and is also provided with
Thawing the intermediate mesenchymal lineage precursor or stem cell preparation in the cryopreservation process and expanding the intermediate mesenchymal lineage precursor or stem cell preparation in the process by passaging expansion.
In one example, the expanded mesenchymal lineage precursor or stem cell preparation has antigenic and active properties comprising:
less than about 0.75% cd45+ cells;
at least about 95% cd105+ cells;
At least about 95% cd166+ cells.
In one example, the expanded mesenchymal lineage precursor or stem cell preparation is capable of inhibiting IL2Ra expression of PBMCs activated by CD3/CD28 by at least about 30% relative to a control.
In one example, the culture expanded mesenchymal lineage precursor or stem cells are culture expanded for about 4-10 passages, wherein the mesenchymal lineage precursor or stem cells are cryopreserved after at least 2 or 3 passages before further culture expansion. In one example, mesenchymal lineage precursor or stem cells are expanded in culture for at least 1, at least 2, at least 3, at least 4, at least 5 passages, cryopreserved and then further expanded in culture for at least 1, at least 2, at least 3, at least 4, at least 5 passages prior to administration or further cryopreservation.
In one example, the majority of mesenchymal lineage precursor or stem cells in the compositions of the present disclosure have about the same algebra (i.e., they are within about 1 or about 2 or about 3 or about 4 cell doublings of each other). In one example, the average number of cell doublings in a composition of the invention is about 20 to about 25 doublings. In one example, the average number of cell doublings in a composition of the invention is about 9 to about 13 (e.g., about 11 or about 11.2) doublings from the primary culture, plus about 1, about 2, about 3, or about 4 doublings/generation (e.g., about 2.5 doublings/generation). Exemplary average cell-doubling in the compositions of the present invention are any of about 13.5, about 16, about 18.5, about 21, about 23.5, about 26, about 28.5, about 31, about 33.5, and about 36 when produced by about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, and about 10 generations, respectively.
The process of mesenchymal lineage precursor or stem cell isolation and ex vivo expansion can be performed using any apparatus and cell handling methods known in the art. Various culture expansion embodiments of the present disclosure employ steps requiring manipulation of cells, e.g., seeding, feeding, dissociating adherent cultures, or washing steps. Any step in manipulating the cells may damage the cells. Although mesenchymal lineage precursor or stem cells can typically sustain a certain amount of damage during the preparation process, it is preferred to manipulate the cells by a treatment procedure and/or apparatus that adequately performs one or more given steps while minimizing damage to the cells.
In one example, mesenchymal lineage precursor or stem cells are washed in a device comprising a cell source bag, a wash solution bag, a recirculating wash bag, a rotating membrane filter with an inlet and an outlet, a filtrate bag, a mixing zone, a final product bag for washed cells, and appropriate tubing, for example, as described in US 6,251,295, which is incorporated herein by application.
In one example, a mesenchymal lineage precursor or stem cell composition according to the present disclosure is 95% homogeneous with respect to CD105 positive and CD166 positive and CD45 negative. In one example, this homogeneity is amplified ex vivo; i.e., persisting through multiple population doublings. In one example, the composition comprises at least one therapeutic dose of mesenchymal lineage precursor or stem cells, and the mesenchymal lineage precursor or stem cells comprise less than about 1.25% cd45+ cells, at least about 95% cd105+ cells, and at least about 95% cd166+ cells. In one example, the homogeneity persists after low temperature storage and thawing, wherein the cells also typically have a viability of about 70% or higher.
In one example, the compositions of the present disclosure comprise mesenchymal lineage precursor or stem cells that express substantial levels of TNF-R1, e.g., greater than 13pg TNF-R1/million mesenchymal lineage precursor or stem cells. In one example, the phenotype is stable during ex vivo amplification and low temperature storage. In one example, TNF-R1 expression levels in the range of about 13 to about 179pg (e.g., about 13pg to about 44 pg)/million mesenchymal lineage precursor or stem cells are correlated with a desired therapeutic potential that also persists throughout ex vivo expansion and cryopreservation.
In one example, the expanded mesenchymal lineage precursor or stem cells are cultured to express tumor necrosis factor receptor 1 (TNF-R1) in an amount of at least 110 pg/ml. For example, mesenchymal lineage precursor or stem cells can express TNF-R1 in an amount of at least 150pg/m1, or at least 200pg/ml, or at least 250pg/ml, or at least 300pg/ml, or at least 320pg/ml, or at least 330pg/ml, or at least 340pg/ml, or at least 350 pg/ml.
In one example, the mesenchymal lineage precursor or stem cells are at least 13pg/10 6 The individual cells expressed TNF-R1. For example, mesenchymal lineage precursor or stem cells are at least 15pg/10 6 Individual cells, or at least 20pg/10 6 Individual cells, or at least 25pg/10 6 Individual cells, or at least 30pg/10 6 Individual cells, or at least 35pg/10 6 Individual cells, or at least 40pg/10 6 Individual cells, or at least 45pg/10 6 Individual cells, or at least 50pg/10 6 The individual cells expressed TNF-R1.
In another example, a mesenchymal lineage precursor or stem cell disclosed herein inhibits IL-2rα expression on a T cell. In one example, the mesenchymal lineage precursor or stem cells are capable of inhibiting IL-2rα expression by at least about 30%, alternatively at least about 35%, alternatively at least about 40%, alternatively at least about 45%, alternatively at least about 50%, alternatively at least about 55%, alternatively at least about 60.
In one example, the compositions of the present disclosure comprise at least one therapeutic dose of mesenchymal lineage precursor or stem cells, which can comprise at least about 10000 ten thousand cells or about 12500 ten thousand cells, for example.
Modification of cells
In one example, the mesenchymal lineage precursor or stem cells of the present disclosure can be altered in such a way that: after administration, lysis of the cells is inhibited. The alteration of the antigen may induce immune anergy or tolerance, thereby preventing the induction of an immune response effector phase (e.g., production of cytotoxic T cells, antibody production, etc.) that ultimately results in rejection of the foreign cells in a normal immune response. Antigens that can be altered to achieve this goal include, for example, MHC class I antigens, MHC class II antigens, LFA-3, and ICAM-1.
Mesenchymal lineage precursor or stem cells can also be genetically modified to express proteins important for differentiation and/or maintenance of striated skeletal muscle cells. Exemplary proteins include growth factors (TGF-beta, insulin-like growth factor 1 (IGF-1), FGF), myogenic factors (e.g., myoD, myogenin, myogenic factor 5 (Myf 5), myogenic Regulatory Factor (MRF)), transcription factors (e.g., GATA-4), cytokines (e.g., cardiomyodin-1), neuregulin family members (e.g., neuregulin 1, 2, and 3), and homeobox genes (e.g., csx, tinman, and NKx families).
The mesenchymal lineage precursor or stem cells of the present disclosure can also be modified to carry or express an antiviral agent or thrombolytic agent. In one example, the agent is an antiviral drug. In one example, the agent is an anti-influenza drug. In one example, the agent is an anti-SARS-CoV (e.g., SARS-Cov 2). An exemplary agent is adefovir. In one example, the agent is a thrombolytic drug. Examples of thrombolytic agents include Eminase (anipase), retavase (reteplase), streptase (streptokinase, carbikinase). In one example, the thrombolytic agent is heparin.
The mesenchymal precursor cells or stem cells of the present disclosure may be modified to carry antiviral or thrombolytic agents by culturing the cells with an agent for a period of time and under conditions sufficient to allow the agent to be taken up by the cells. In one example, an antiviral or thrombolytic agent is added to a culture medium of mesenchymal lineage precursor or stem cells disclosed herein. For example, mesenchymal lineage precursor or stem cells disclosed herein can be culture expanded in a medium comprising an antiviral or thrombolytic agent.
In another example, the antiviral or thrombolytic agent is a peptide. In one example, mesenchymal lineage precursor or stem cells are genetically modified to express an antiviral or thrombolytic peptide or nucleic acid encoding the same. In one example, the mesenchymal lineage precursor or stem cells are modified by contacting in vitro with a viral vector. For example, the virus may be added to the cell culture medium. Genetically modified non-viral methods may also be employed. Examples include plasmid transfer and applications for targeted gene integration by using integrase or transposase technology, liposome or protein transduction domain mediated delivery, and physical methods (e.g., electroporation).
The efficiency of genetic modification is rarely 100% and it is often necessary to enrich cell populations that have been successfully modified. In one example, modified cells can be enriched by exploiting the functional characteristics of the new genotype. One exemplary method of enriching for modified cells is forward selection using selectable or screenable marker genes. "marker gene" refers to a gene that confers a different phenotype to cells expressing the marker gene and thus allows such transformed cells to be distinguished from cells without the marker. Selectable marker genes confer a trait that is "selectable" based on resistance to a selection agent (e.g., an antibiotic). The screenable marker gene (or reporter gene) confers a trait that can be identified by observation or testing, i.e., by "screening" (e.g., β -glucuronidase, luciferase, GFP, or other enzymatic activities that are not present in untransformed cells). In one example, genetically modified mesenchymal lineage precursor or stem cells are selected based on resistance to a drug (e.g., neomycin), or colorimetric selection is performed based on lacZ expression.
Compositions of the present disclosure
In one example of the present disclosure, mesenchymal lineage precursor or stem cells and/or progeny thereof and/or soluble factors derived therefrom are administered in the form of a composition. In one example, such compositions comprise a pharmaceutically acceptable carrier and/or excipient. Thus, in one example, the compositions of the present disclosure may comprise culture expanded mesenchymal lineage precursor or stem cells.
The terms "carrier" and "excipient" refer to compositions of matter conventionally used in the art to facilitate storage, administration, and/or biological activity of an active compound (see, e.g., remington's Pharmaceutical Sciences, 16 th edition, mac Publishing Company (1980). A carrier may also reduce any undesirable side effects of an active compound.
Suitable carriers for the present disclosure include those conventionally used, such as water, saline, aqueous dextrose, lactose, ringer's solution, buffer solutions, hyaluronic acid and glycols are exemplary liquid carriers, particularly (when isotonic) for use in solutions. Suitable pharmaceutical carriers and excipients include starch, cellulose, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, magnesium stearate, sodium stearate, glycerol monostearate, sodium chloride, glycerol, propylene glycol, water, ethanol and the like.
In another example, the carrier is a medium composition, such as a cell grown or suspended therein. For example, such a medium composition does not cause any side effects in the subject to which it is administered.
Exemplary carriers and excipients do not adversely affect the viability of the cells and/or the ability of the cells to reduce, prevent or delay metabolic syndrome and/or obesity.
In one example, the carrier or excipient provides buffer activity to maintain the cells and/or soluble factors at a suitable pH to exert biological activity, e.g., the carrier or excipient is Phosphate Buffered Saline (PBS). PBS represents an attractive carrier or excipient because it minimally interacts with and allows for rapid release of cells and factors, in which case the compositions of the present disclosure may be produced as a liquid for direct application to the blood stream or into tissue or surrounding or adjacent areas of tissue, such as by injection.
The mesenchymal lineage precursor or stem cells and/or progeny thereof and/or soluble factors derived therefrom can also be incorporated or embedded into a receptor compatible scaffold, and the scaffold degraded to products harmless to the receptor. These scaffolds provide support and protection for cells to be transplanted into a recipient subject. Natural and/or synthetic biodegradable scaffolds are examples of such scaffolds.
A variety of different stents may be successfully used in the practice of the present disclosure. Exemplary stents include, but are not limited to, biological, degradable stents. Natural biodegradable scaffolds include collagen, fibronectin and laminin scaffolds. Suitable synthetic materials for cell transplantation scaffolds should be capable of supporting a wide range of cell growth and cell functions. Such stents may also be resorbable. Suitable scaffolds include polyglycolic acid scaffolds (e.g., as described by Vacanti, et al J. Ped. Surg.23:3-91988; cima, et al Biotechnol. Bioeng. 38:1451991; vacanti, et al Plast. Reconstre. Surg. 88:753-91991); or synthetic polymers such as polyanhydrides, polyorthoesters and polylactic acid.
In another example, mesenchymal lineage precursor or stem cells and/or progeny thereof and/or soluble factors derived therefrom can be administered in a gel scaffold (such as Gelfoam from Upjohm Company).
The compositions described herein may be administered alone or as a mixture with other cells. The different types of cells may be mixed with the compositions of the present disclosure immediately or shortly before administration, or they may be co-cultured together for a period of time prior to administration.
In one example, the composition or total dose comprises an effective or therapeutically effective or prophylactically effective amount of mesenchymal lineage precursor or stem cells and/or progeny thereof and/or soluble factors derived therefrom. For example, the total dose contains more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg). In another example, the total dose comprises more than 600 tens of thousands of cells// kg). In one example, the total dose comprises more than 500 tens of thousands of cells/kg. In another example, more than 600 ten thousand cells/kg are administered to the subject. In another example, more than 800 ten thousand cells/kg are administered to the subject.
In one example, more than 400 ten thousand MLPSCs/kg are administered to a subject by at least 2 to 3 doses. In one example, the subject receives at least 3 doses. In another example, the subject receives more than 400 ten thousand MLPSCs/kg within 5 to 9 days of administration of the first dose. In another example, the subject receives more than 400 ten thousand MLPSCs/kg within 7 days of administration of the first dose. In these examples, the dose is contained in 1X 10 8 And 2.5X10 8 Individual cells/dose. For example, the dosage comprises 1.6X10 0 Individual cells. In another example, the dose comprises 200 tens of thousands of cells/kg.
In one example, the composition comprises greater than 5.00 x 10 6 Each living cell/mL. In another example, the composition comprises greater than 5.50X10 6 Each living cell/mL. In another example, the composition comprises greater than 6.00 x 10 6 Each living cell/mL. In another example, the composition comprises greater than 6.50X10 6 Each living cell/mL. In another example, the composition comprises greater than 6.68X10 6 Each living cell/mL.
In one example, the mesenchymal lineage precursor or stem cells comprise at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99% of the cell population of the composition.
The compositions of the present disclosure may be cryopreserved. Cryopreservation of mesenchymal lineage precursor or stem cells can be performed using slow freezing methods or fast freezing protocols known in the art. Preferably, the cryopreservation method maintains similar phenotypes, cell surface markers and growth rates of cryopreserved cells compared to unfrozen cells.
The cryopreserved composition may comprise a cryopreservation solution. The pH of the cryopreservation solution is generally from 6.5 to 8, preferably 7.4.
The cryopreservation solution may comprise a sterile, pyrogen-free isotonic solution, such as, for example, P1asmaLyte A TM 。100mL P1asmaLyte A TM Contains 526mg sodium chloride, USP (NaCl); 502mg sodium gluconate (C) 6 H 11 NaO 7 ) The method comprises the steps of carrying out a first treatment on the surface of the 368mg sodium acetate trihydrate, USP (C 2 H 3 NaO 2 ·3H 2 O); 37mg potassium chloride, USP (KCl); and 30mg magnesium chloride, USP (MgCl 2 ·6H 2 O). It does not contain an antimicrobial agent. The pH was adjusted with sodium hydroxide. The pH was 7.4 (6.5 to 8.0).
The cryopreservation solution may comprise Profreize TM . The cryopreservation solution may alternatively or alternatively comprise a culture medium, for example αmem.
To facilitate freezing, cryoprotectants such as, for example, dimethyl sulfoxide (DMSO) are typically added to the cryopreservation solution. Ideally, cryoprotectants should be non-toxic, non-antigenic, chemically inert to cells and patients, provide high survival rates after thawing, and be transplanted without washing. However, the most commonly used cryoprotectant DMSO shows some cytotoxicity. Hydroxyethyl starch (HES) may be used as an alternative or in combination with DMSO to reduce cytotoxicity of the cryopreservation solution.
The cryopreservation solution may comprise one or more of DMSO, hydroxyethyl starch, human serum components, and other protein fillers. In one example, the cryopreserved solution comprises about 5% Human Serum Albumin (HSA) and about 10% dmso. The cryopreservation solution may further comprise one or more of methylcellulose, polyvinylpyrrolidone (PVP), and trehalose.
In one implementationIn the protocol, cells were suspended in 42.5% Profreze TM 50% alpha MEM/7.5% DMSO, and cooled in a controlled speed refrigerator.
The cryopreserved composition may be thawed and applied directly to a subject or added to another solution, e.g., a solution comprising HA. Alternatively, the cryopreserved composition may be thawed prior to administration and the mesenchymal lineage precursor or stem cells resuspended in an alternative carrier.
In one example, the cell composition of the present disclosure may comprise Plasma-Lyte a, dimethyl sulfoxide (DMSO), and Human Serum Albumin (HSA). For example, the compositions of the present disclosure may comprise a Plasma-Lyte a (70%), DMSO (10%), HSA (25%) solution, HSA solution comprising 5% HSA and 15% buffer.
In one example, the compositions described herein can be administered as a single dose.
In some examples, the compositions described herein may be administered in a variety of doses. For example, at least 2, at least 3, at least 4 doses. In other examples, the compositions described herein may be administered by at least 5, at least 6, at least 7, at least 8, at least 9, at least 10 doses.
In one example, the mesenchymal lineage precursor or stem cells can be culture expanded prior to administration to the subject. Various methods of cell culture are known in the art. In one example, mesenchymal lineage precursor or stem cells are expanded in culture for about 4-10 passages. In one example, the mesenchymal lineage precursor or stem cells are expanded in culture for at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10 passages. In one example, mesenchymal lineage precursor or stem cells are expanded in culture for at least 5 passages. In these examples, the stem cells may be culture expanded prior to cryopreservation.
In one example, prior to administration, mesenchymal lineage precursor or stem cells are culture expanded in serum-free medium.
In some examples, the cells are contained in a chamber that does not allow the cells to exit into the circulatory system of the subject, but allows factors secreted by the cells to enter into the circulatory system. In this way, the soluble factor may be administered to the subject by allowing the cell to secrete the factor into the circulation of the subject. Such a chamber may likewise be implanted at a site in a subject to increase the local level of soluble factors.
In one example, mesenchymal lineage precursor or stem cells can be administered systemically. In one example, mesenchymal lineage precursor or stem cells can be administered to the airway of a subject. In one example, mesenchymal lineage precursor or stem cells can be administered to the lung of a subject. In another example, the compositions of the present disclosure may be administered intravenously. In another example, the composition is administered intravenously and administered to the airway of the subject.
In one example, mesenchymal lineage precursor or stem cells are administered twice weekly. In one example, mesenchymal lineage precursor or stem cells can be administered once a month. In one example, two doses of mesenchymal lineage precursor or stem cells are administered once per week within two weeks. In another example, two doses of mesenchymal lineage precursor or stem cells are administered weekly every two weeks. In another example, four doses of mesenchymal lineage precursor or stem cells are administered within two weeks, followed by a subsequent dose administered monthly. In one example, two doses of mesenchymal lineage precursor or stem cells are administered weekly every two weeks, followed by a subsequent dose administered monthly. In one example, four doses are administered monthly.
In one example, the compositions of the present disclosure comprise a "clinically proven effective" amount of MLPSC. In one example, the compositions of the present disclosure comprise a "clinically proven effective" amount of MSCs.
Those skilled in the art will appreciate that many changes and/or modifications may be made to the above-described embodiments without departing from the broad general scope of the present disclosure. The present embodiments are, therefore, to be considered in all respects as illustrative and not restrictive.
The following specific examples are to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever. Without further elaboration, it is believed that one skilled in the art can, based on the description herein, utilize the present invention to its fullest extent.
Examples
Ex vivo culturing of expanded adult allogeneic bone marrow derived Mesenchymal Stem Cells (MSC) for the treatment of acute respiratory distress syndrome (ARRDS)
Composition and method for producing the same
The composition consists of culture expanded mesenchymal stem cells (ceMSCs) isolated from bone marrow of healthy adult donors. The final composition contained ceMSC formulated in Plasma-Lyte A, dimethyl sulfoxide (DMSO) and Human Serum Albumin (HSA).
Target object
To determine:
security-security
Improvements in survival and respiratory function
-a change in biomarker level.
A subject
Patients were characterized as suffering from moderate covd-19 associated ARDS and receiving mesenchymal stem cells (intravenous; 200 ten thousand (2 x 10) 6 ) Fixed dosing regimen within 3-5 days per kilogram x 2 dose) or control therapy. 125 patients were aged < 65 years (i.e., < 65 years of Intent To Treat (ITT) population; table 1). 58 < 65 years of ITT populations received cell therapy and 67 received control therapy. Some < 65 years of ITT patients were identified as suffering from mild ARRDS and/or low levels of inflammation characterized by circulating CRP < 4. These patients were excluded from the protocol (PP) patient population. Thus, PP populations were reduced to 89 (from 125), 38 of which received cell therapy and 51 received control therapy. Furthermore, the PP population represents the study population that always had the most severe disease, as they all had moderate to severe ARDS.97 patients were aged > 65 years (i.e., > 65 years of Intent To Treat (ITT) population; table 1). 54, the ITT population > 65 years old received cell therapy, and 43 received control therapy. Some ITT patient rows > 65 years oldExcept for the analysis, the total number of patients was reduced to 94 persons (referred to as corrected intent-to-treat population (mtt)).
Table 1: baseline summary data: patients with predetermined ages < 65 and > 65 were intended to be treated.
Analysis
Analysis of the treated patients surprisingly revealed that cell therapy provided significant protection from death in patients aged less than 65 on day 60 (FIGS. 1 and 2;A) relative to patients aged greater than 65 (FIGS. 1 and 2; B). Notably, protection from death on day 60 was evident in patients with Intent To Treat (ITT) patients (38.3% reduction in death compared to control; p= 0.0484; fig. 1) and in patients on protocol (PP) patients (43.3% reduction in death compared to control; p=0.0285; fig. 2) with ages less than 65. As also shown in fig. 1 and 2, a significant reduction in mortality was observed in PP patients compared to ITT patients. This is an unexpected result, since PP patients always suffer from more severe disease than ITT patients.
Further analysis of the treated patients revealed a durable improvement in respiratory function in patients < 65 years from day 7 to day 30 (FIG. 3; improvement measures are regression and/or improvement of ARDS 7, 14, 21 and 30 days after randomization as defined by Berlin standards). Improved respiratory function was also observed in treated > 65 year old patients (fig. 3). However, in contrast to < 65 years old patients, improved respiratory function did not persist from day 14 on in treated, > 65 years old subjects (fig. 3). These findings support the need for higher accumulation or longer dosing (i.e., over 400 tens of thousands of cells/kg) in patients > 65 years of age.
Biomarker analysis
In 107 treated patients and 106 controls (treated < 65 years, n=55; control < 65 years, n=65; treated > 65 years, n=52; control > 65 years, n=41), at baseline and in the treatment with cell therapy (200 ten thousand (2×10) 6 ) Individual cells/kg x 2 doses, fixed dosing regimen over 3-5 days). In patients < 65 years old, cell therapy significantly reduced CRP levels at days 3, 7 and 14 (fig. 4A) and ferritin levels at days 7 and 14 (fig. 4B). However, similar results were not observed in patients > 65, further supporting the above referenced requirements for higher doses or longer dosing in these patients (fig. 4A and 4B). Cell therapy prevented a significant increase in D-dimer levels in all treated patients (fig. 4C).
Comparison of inflammatory biomarker levels between treated subjects < 65 years and > 65 years provides unique opportunities to identify measurable criteria for selecting patients for treatment with higher doses or longer doses (i.e., over 400 tens of thousands of cells/kg). Surprisingly, as shown in figures 5 and 6, an age > 65 years appears to be associated with an increase in inflammatory activity at baseline, whether or not the patient is taking corticosteroids. Indeed, analysis of all patients taking corticosteroids at baseline showed > 5-fold higher inflammatory activity (cytokines/chemokines) in patients > 65 years of age than in patients < 65 years of age.
Figure 5 shows that patients older than 65 years old have higher baseline inflammatory cytokine/chemokine levels than patients < 65, particularly:
(i) CCR 2-binding chemokines (including CXCL10/IP10 and CXCL 9) and CXCR 3-binding chemokines (including CCL2, CCL3 and CCL7/MCP 3). This group of chemokines indicates increased neutrophil and macrophage influx into the lung.
(ii) IL-6 and IL-8 indicate increased macrophage inflammation and increased neutrophil migration to the lung.
(ii) CCL19 and IL-2, which indicate T cell activation/proliferation and apoptotic death.
Other inflammatory biomarkers that were increased in patients > 65 years old include TGF-alpha, TNF, CXCL8, G-CSF/CSF-3, and IL17C (FIGS. 5 and 6).
Comparison of inflammatory pathways between < 65 years old patients and > 65 years old patients reveals further support for the above referenced requirements for higher doses or longer administrations in > 65 years old patients and/or patients with biomarker-defined increased inflammatory activity at baseline. Patients aged 65 express the same inflammatory pathway as patients > 65, albeit at lower baseline levels. In < 65 patients, cell therapy reduces inflammatory markers, in particular: is associated with a severity of covd-19 ARDS in < 65 patients, in particular:
-CCR2 and CXCR3 binding chemokines may lead to reduced neutrophil and macrophage influx into the lung;
-IL-6/IL-8/TNF/IL-18, indicating IL-18 driven inflammatory body reduction, and macrophage activation/neutrophil homing to lung reduction;
-CCL19 decrease and IL-4/IL-13/GM-CSF indicates reduced T cell influx and activation.
Since patients are involved in the same pathways regardless of age, the data shows that these pathways remain over-expressed at higher levels in patients > 65 years old, suggesting that cell therapy would also be effective in elderly patients (or patients with higher baseline levels of inflammation) if used with a higher dosing regimen.
To sum up:
inflammatory pathways were down-regulated in treated < 65 year old patients (fig. 5 and 6), and this corresponds to improved prognosis (fig. 1 and 2) and persistent improvement of respiratory function (fig. 3);
in treated patients > 65 years old, improved respiratory function was observed on day 7, but failed to persist (fig. 3); subsequent biomarker analysis revealed that these patients had higher baseline levels of inflammation (figures 5 and 6);
biomarker analysis revealed similar inflammatory pathways were involved in ARDS patients regardless of age (< 65 vs > 65 years). Thus, the higher baseline level of inflammation observed in patients > 65 years old, as well as the initial improvement in lung function observed on day 7 in these patients, suggests that improved results may be achieved with higher doses of stem cells or longer administration (e.g., comparable to those of patients < 65 years old).
In particular, the data indicate that administration is included in excess of 2X 10 6 A dosing regimen of individual cells/kg x 2 doses (i.e., 400 tens of thousands of cells/kg) would be therapeutically effective in patients with an increased baseline level of inflammatory biomarkers at baseline and/or > 65 years old.
Safety of
No adverse events associated with infusion occurred.
Those skilled in the art will appreciate that various changes and/or modifications may be made to the invention as shown in the specific embodiments without departing from the spirit or scope of the invention as broadly described. The present embodiments are, therefore, to be considered in all respects as illustrative and not restrictive.
All publications discussed above are incorporated herein in their entirety.
The present application claims priority from AU2021901214 submitted at month 2021, 23, AU2021902180 submitted at month 2021, 7, 15, 2022, 2, 9, 2022900260, and AU2022900372 submitted at month 2022, 18, the disclosures of which are incorporated herein by reference.
Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is solely for the purpose of providing a context for the present invention. It is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present invention as it existed before the priority date of each claim of this application.
Claims (52)
1. A method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising selecting a subject with ARDS greater than or equal to 65 years of age, administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg).
2. The method of claim 1, wherein the subject has an increased biomarker of inflammation that is indicative of:
(a) Increased flow of neutrophils and macrophages into the lung;
(b) Macrophage inflammation and neutrophil migration to the lung are increased; and/or the number of the groups of groups,
(c) T cell activation/proliferation and apoptotic death.
3. A method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising selecting a subject with ARDS having an increased inflammatory biomarker indicative of:
(a) Increased flow of neutrophils and macrophages into the lung;
(b) Macrophage inflammation and neutrophil migration to the lung are increased; and/or the number of the groups of groups,
(c) T cell activation/proliferation and apoptotic death,
and administering to the subject more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kilogram body weight (cells/kg).
4. A method according to any one of claims 1 to 3, wherein the subject is ventilator dependent.
5. The method of any one of claims 2 to 4, wherein the inflammatory biomarker indicative of increased neutrophil and macrophage influx into the lung is CCR2 or CXCR3 binding chemokine.
6. The method of claim 5, wherein the one or more CCR 2-binding chemokines is CCL2, CCL3, or CCL7, preferably CCL2.
7. The method of claim 5, wherein the CXCR 3-binding chemokine is CXCL10 or CXCL9, preferably CXCL10.
8. The method of any one of claims 2 to 7, wherein the inflammatory biomarker indicative of increased macrophage inflammation and increased neutrophil migration to the lung is IL-6 or IL-8.
9. The method of any one of claims 2 to 8, wherein the inflammatory biomarker indicative of T cell activation/proliferation and apoptotic death is CCL19 or IL-2.
10. The method of any one of claims 1 to 9, wherein the subject has an increased level of one or more inflammatory biomarkers relative to a subject less than 65 years old.
11. The method of claim 10, wherein the level of one or more inflammatory biomarkers in the subject is increased by at least 2-fold relative to a subject less than 65 years old.
12. The method of claim 10, wherein the level of one or more inflammatory biomarkers in the subject is increased by at least 5-fold relative to a subject less than 65 years old.
13. The method of any one of claims 3 to 12, wherein the subject is greater than or equal to 65 years old.
14. The method of any one of claims 1 to 13, wherein the subject has sustained CRP levels after administration of a first dose of MLPSC comprising less than 400 ten thousand MLPSCs per kilogram body weight (cells/kg).
15. The method of claim 14, wherein the subject has sustained CRP levels 3 days after administration of the first dose of MLPSC.
16. The method of any one of claims 4 to 15, wherein the subject is disconnected from the ventilator after treatment.
17. The method of claim 16, wherein the subject is out of ventilator within 60 days of treatment.
18. A method of treating or preventing Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising:
-determining or having determined the level of one or more inflammatory biomarkers of the subject, said inflammatory biomarkers being selected from the list comprising: (i) CXCR3 binds to chemokines, preferably CXCL10 and/or CXCL9; (ii) CCR 2-binding chemokines, preferably CCL2, CCL3 and/or CCL7; (iii) IL-6; (v) IL-8; (vi) CCL19; (vii) IL-2; and/or (viii) CRP;
-selecting a subject, said subject:
o is greater than or equal to 65 years old; and/or the number of the groups of groups,
increased levels of one or more inflammatory biomarkers relative to a subject less than 65 years old; and/or the number of the groups of groups,
3 days after administration of a first dose of mesenchymal lineage precursor or stem cells (MLPSC) comprising less than 400 ten thousand MLPSC per kg body weight (cells/kg), with sustained CRP levels; and
-administering more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC) per kg body weight (cells/kg) to the subject.
19. The method of any one of claims 1 to 18, wherein treatment reduces the level of at least one inflammatory biomarker relative to baseline, wherein the at least one inflammatory biomarker is indicative of:
(a) Neutrophil and macrophage influx into the lungs is reduced;
(b) Reduction of inflammatory body;
(c) Macrophage activation and decreased neutrophil migration to the lung;
(d) Reduced T cell influx and activation; or (b)
(e) Circulating biomarkers of macrophage and neutrophil inflammation are reduced.
20. The method of claim 19, wherein the one or more inflammatory biomarkers are one or more of:
-CXCR 3-binding chemokines, preferably CXCL10 and/or CXCL9;
-CCR 2-binding chemokines, preferably CCL2, CCL3 and/or CCL7;
-IL-6;
-IL-8;
-TNF;
-IL-18;
-CCL19;
-IL-4;
-IL-13;
-GM-CSF;
-CRP; or (b)
Ferritin.
21. The method of any one of claims 1 to 20, wherein the treatment reduces CRP and/or ferritin levels within 3 to 14 days of administration of more than 400 ten thousand MLPSCs per kilogram body weight (cells/kg).
22. The method of any one of claims 1-21, wherein treatment improves respiratory function of the subject.
23. The method of claim 22, wherein respiratory function defined by berlin standards is improved on day 14 and/or day 21.
24. The method of any one of claims 1-23, wherein more than 500 ten thousand MLPSCs/kg, more than 600 ten thousand MLPSCs/kg, or more than 800 ten thousand MLPSCs/kg are administered to the subject.
25. The method of any one of claims 1 to 24, wherein the greater than 400 ten thousand MLPSCs/kg are administered by at least 2 to 3 doses.
26. The method of claim 25, wherein the subject receives more than 400 ten thousand MLPSCs per kilogram of body weight (cells/kg) within 5 to 9 days of administration of the first dose.
27. The method of claim 25, wherein the subject receives more than 400 ten thousand MLPSCs per kilogram of body weight (cells/kg) within 7 days of administration of the first dose.
28. The method of any one of claims 25 to 27, comprising administering 1 x 10 8 Sum 2.5×10 8 MLPSC/dose.
29. The method of any one of claims 25 to 27, comprising administering about 1.6x10 8 MLPSC/dose.
30. The method of any one of claims 1-29, wherein the subject is also taking a corticosteroid.
31. The method of any one of claims 1 to 29, further comprising a corticosteroid.
32. The method of any one of claims 30 or 31, wherein the corticosteroid is dexamethasone.
33. The method of any one of claims 1 to 32, wherein the ARDS is moderate or severe.
34. The method of any one of claims 1 to 33, wherein the ARDS is caused by a viral infection such as rhinovirus, influenza virus, respiratory Syncytial Virus (RSV) or coronavirus.
35. The method of claim 34, wherein the viral infection is caused by a coronavirus.
36. The method of claim 35, wherein the coronavirus is severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV), or covd-19.
37. The method of any one of claims 1 to 36, wherein the MLPSC has been cryopreserved and thawed.
38. The method of any one of claims 1 to 37, wherein the MLPSCs are culture expanded from an intermediate cryopreserved MLPSC population.
39. The method of claim 38, wherein the MLPSC is culture amplified for at least about 5 generations.
40. The method of any one of claims 1-39, wherein the MLPSC expresses at least 13pg TNFR1/million MLPSCs.
41. The method of any one of claims 1-40, wherein the MLPSC expresses about 13pg to about 44pg TNFR1/million MLPSCs.
42. The method of any one of claims 38 to 41, wherein the culture expansion comprises at least 20 or 30 population doublings.
43. The method of any one of claims 1-42, wherein the MLPSC is a Mesenchymal Stem Cell (MSC).
44. The method of any one of claims 1 to 43, wherein the MLPSC is allogeneic.
45. The method of any one of claims 1 to 44, wherein the MLPSC is modified to carry or express an antiviral drug or thrombolytic agent.
46. The method of any one of claims 1-45, wherein the MLPSC is administered in one or more compositions comprising Plasma-Lyte a, dimethyl sulfoxide (DMSO), human Serum Albumin (HSA).
47. The method of claim 46, wherein the composition comprises a solution of Plasma-Lyte a (70%), DMSO (10%), HSA (25%), the HSA solution comprising 5% HSA and 15% buffer.
48. The method of claim 46 or 47, wherein the composition comprises more than 6.68 x 10 6 Each living cell/mL.
49. A method of treating Acute Respiratory Distress Syndrome (ARDS) in a human subject in need thereof, the method comprising:
-administering 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kg body weight (cells/kg) to the subject;
-determining or having determined the subject level of one or more biomarkers selected from the group consisting of: CXCL10, CXCL9, CCL2, CCL3, CCL7, IL-6, IL-8, CCL19, IL-2, ferritin and/or CRP;
-administering to the subject an additional dose of MLPSC if the level of one or more biomarkers does not decrease from baseline, wherein after the additional dose more than 400 ten thousand mesenchymal lineage precursor or stem cells (MLPSC)/kilogram body weight (cells/kg) have been administered to the subject.
50. The method of claim 49, wherein the biomarker is CRP and/or ferritin.
51. The method of claim 49, wherein CRP and/or ferritin levels of the subject are determined relative to baseline on day 7 and/or day 14.
52. The method of any one of claims 49 to 51, wherein the additional dose comprises 200 ten thousand MLPSCs per kilogram body weight (cells/kg).
Applications Claiming Priority (6)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
AU2021901214 | 2021-04-23 | ||
AU2021902180 | 2021-07-15 | ||
AU2022900260 | 2022-02-09 | ||
AU2022900372A AU2022900372A0 (en) | 2022-02-18 | Method for treating acute respiratory distress syndrome (ARDS) using mesenchymal lineage precursor or stem cells | |
AU2022900372 | 2022-02-18 | ||
PCT/IB2022/053762 WO2022224206A1 (en) | 2021-04-23 | 2022-04-22 | Method for treating acute respiratory distress syndrome (ards) in specific patients using mesenchymal lineage precursor or stem cells |
Publications (1)
Publication Number | Publication Date |
---|---|
CN117715649A true CN117715649A (en) | 2024-03-15 |
Family
ID=89295837
Family Applications (2)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202280034316.5A Pending CN117715649A (en) | 2021-04-23 | 2022-04-22 | Methods of treating Acute Respiratory Distress Syndrome (ARDS) in a particular patient using mesenchymal lineage precursor or stem cells |
CN202280035460.0A Pending CN117337184A (en) | 2021-04-23 | 2022-04-22 | Methods of treating Acute Respiratory Distress Syndrome (ARDS) using mesenchymal lineage precursor or stem cells |
Family Applications After (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202280035460.0A Pending CN117337184A (en) | 2021-04-23 | 2022-04-22 | Methods of treating Acute Respiratory Distress Syndrome (ARDS) using mesenchymal lineage precursor or stem cells |
Country Status (1)
Country | Link |
---|---|
CN (2) | CN117715649A (en) |
-
2022
- 2022-04-22 CN CN202280034316.5A patent/CN117715649A/en active Pending
- 2022-04-22 CN CN202280035460.0A patent/CN117337184A/en active Pending
Also Published As
Publication number | Publication date |
---|---|
CN117337184A (en) | 2024-01-02 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
AU2021202391B2 (en) | Cells with increased immuno-regulatory properties and methods for their use and manufacture | |
JP6755850B2 (en) | Use of mesenchymal stem cells | |
KR20220164741A (en) | Method for treating hyperinflammation using mesenchymal progenitor cells or stem cells | |
CN116057173A (en) | Composition comprising mesenchymal precursor cells or stem cells and uses thereof | |
US20230293589A1 (en) | Method for treating inflammatory lung diseases using mesenchymal lineage precursor or stem cells | |
US20240197787A1 (en) | Method for treating acute respiratory distress syndrome (ards) in specific patients using mesenchymal lineage precursor or stem cells | |
CN117715649A (en) | Methods of treating Acute Respiratory Distress Syndrome (ARDS) in a particular patient using mesenchymal lineage precursor or stem cells | |
CN115361960A (en) | Methods for treating chronic graft versus host disease | |
KR20220152240A (en) | Methods of treating inflammatory bowel disease II | |
WO2023119239A1 (en) | Method of treating severe graft versus host disease | |
WO2024121819A1 (en) | Method of treating inflammation with cellular compositions | |
EP4264275A2 (en) | Method of treating progressive heart failure in subjects with class ii heart failure | |
AU2021403025A1 (en) | Method of treating progressive heart failure in subjects with class ii heart failure | |
WO2023092043A1 (en) | Method of treating progressive heart failure in subjects at high risk of poor outcomes | |
CN116829162A (en) | Methods of treating progressive heart failure in a subject suffering from grade II heart failure | |
CN115361957A (en) | Method for treating inflammatory bowel disease I |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination |