US20230226129A1 - Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy - Google Patents
Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy Download PDFInfo
- Publication number
- US20230226129A1 US20230226129A1 US18/159,482 US202318159482A US2023226129A1 US 20230226129 A1 US20230226129 A1 US 20230226129A1 US 202318159482 A US202318159482 A US 202318159482A US 2023226129 A1 US2023226129 A1 US 2023226129A1
- Authority
- US
- United States
- Prior art keywords
- bacteria
- bacteroides
- mice
- myocarditis
- cells
- 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.)
- Abandoned
Links
- 230000001580 bacterial effect Effects 0.000 title description 82
- 230000000112 colonic effect Effects 0.000 title description 24
- 230000002757 inflammatory effect Effects 0.000 title description 23
- 208000031229 Cardiomyopathies Diseases 0.000 title description 15
- 231100000518 lethal Toxicity 0.000 title description 10
- 230000001665 lethal effect Effects 0.000 title description 10
- 230000008030 elimination Effects 0.000 title description 9
- 238000003379 elimination reaction Methods 0.000 title description 9
- 208000009525 Myocarditis Diseases 0.000 claims abstract description 98
- 238000000034 method Methods 0.000 claims abstract description 79
- 241000606125 Bacteroides Species 0.000 claims abstract description 66
- 241001148536 Bacteroides sp. Species 0.000 claims abstract description 51
- 238000003745 diagnosis Methods 0.000 claims abstract description 9
- 230000000670 limiting effect Effects 0.000 claims abstract description 9
- 241000894006 Bacteria Species 0.000 claims description 157
- 239000003242 anti bacterial agent Substances 0.000 claims description 60
- 241000606123 Bacteroides thetaiotaomicron Species 0.000 claims description 44
- 229940088710 antibiotic agent Drugs 0.000 claims description 44
- 230000003115 biocidal effect Effects 0.000 claims description 44
- 101710163270 Nuclease Proteins 0.000 claims description 33
- 108010062877 Bacteriocins Proteins 0.000 claims description 32
- 108010017070 Zinc Finger Nucleases Proteins 0.000 claims description 32
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 claims description 28
- KDXKERNSBIXSRK-UHFFFAOYSA-N lysine Chemical compound NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 claims description 25
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 claims description 24
- PJSFRIWCGOHTNF-UHFFFAOYSA-N Sulphormetoxin Chemical compound COC1=NC=NC(NS(=O)(=O)C=2C=CC(N)=CC=2)=C1OC PJSFRIWCGOHTNF-UHFFFAOYSA-N 0.000 claims description 18
- 229960000282 metronidazole Drugs 0.000 claims description 18
- VAOCPAMSLUNLGC-UHFFFAOYSA-N metronidazole Chemical compound CC1=NC=C([N+]([O-])=O)N1CCO VAOCPAMSLUNLGC-UHFFFAOYSA-N 0.000 claims description 18
- MYPYJXKWCTUITO-UHFFFAOYSA-N vancomycin Natural products O1C(C(=C2)Cl)=CC=C2C(O)C(C(NC(C2=CC(O)=CC(O)=C2C=2C(O)=CC=C3C=2)C(O)=O)=O)NC(=O)C3NC(=O)C2NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(CC(C)C)NC)C(O)C(C=C3Cl)=CC=C3OC3=CC2=CC1=C3OC1OC(CO)C(O)C(O)C1OC1CC(C)(N)C(O)C(C)O1 MYPYJXKWCTUITO-UHFFFAOYSA-N 0.000 claims description 17
- 108010059993 Vancomycin Proteins 0.000 claims description 16
- IEDVJHCEMCRBQM-UHFFFAOYSA-N trimethoprim Chemical compound COC1=C(OC)C(OC)=CC(CC=2C(=NC(N)=NC=2)N)=C1 IEDVJHCEMCRBQM-UHFFFAOYSA-N 0.000 claims description 16
- 229960001082 trimethoprim Drugs 0.000 claims description 16
- 229960003165 vancomycin Drugs 0.000 claims description 16
- MYPYJXKWCTUITO-LYRMYLQWSA-N vancomycin Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C2C=C3C=C1OC1=CC=C(C=C1Cl)[C@@H](O)[C@H](C(N[C@@H](CC(N)=O)C(=O)N[C@H]3C(=O)N[C@H]1C(=O)N[C@H](C(N[C@@H](C3=CC(O)=CC(O)=C3C=3C(O)=CC=C1C=3)C(O)=O)=O)[C@H](O)C1=CC=C(C(=C1)Cl)O2)=O)NC(=O)[C@@H](CC(C)C)NC)[C@H]1C[C@](C)(N)[C@H](O)[C@H](C)O1 MYPYJXKWCTUITO-LYRMYLQWSA-N 0.000 claims description 16
- 102000018120 Recombinases Human genes 0.000 claims description 15
- 108010091086 Recombinases Proteins 0.000 claims description 15
- 238000010459 TALEN Methods 0.000 claims description 14
- 229960005322 streptomycin Drugs 0.000 claims description 14
- 241000956551 Bacteroides faecis Species 0.000 claims description 13
- 229960002227 clindamycin Drugs 0.000 claims description 13
- KDLRVYVGXIQJDK-AWPVFWJPSA-N clindamycin Chemical compound CN1C[C@H](CCC)C[C@H]1C(=O)N[C@H]([C@H](C)Cl)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 KDLRVYVGXIQJDK-AWPVFWJPSA-N 0.000 claims description 13
- 108010043645 Transcription Activator-Like Effector Nucleases Proteins 0.000 claims description 9
- 239000012472 biological sample Substances 0.000 claims description 9
- 239000013068 control sample Substances 0.000 claims description 6
- 239000000203 mixture Substances 0.000 abstract description 83
- 206010056370 Congestive cardiomyopathy Diseases 0.000 abstract description 26
- 201000010046 Dilated cardiomyopathy Diseases 0.000 abstract description 26
- 238000011065 in-situ storage Methods 0.000 abstract description 15
- 241000699670 Mus sp. Species 0.000 description 139
- 108090000623 proteins and genes Proteins 0.000 description 123
- 108090000765 processed proteins & peptides Proteins 0.000 description 85
- 210000004027 cell Anatomy 0.000 description 74
- 210000001744 T-lymphocyte Anatomy 0.000 description 65
- 235000018102 proteins Nutrition 0.000 description 62
- 102000004169 proteins and genes Human genes 0.000 description 62
- 101000716102 Homo sapiens T-cell surface glycoprotein CD4 Proteins 0.000 description 61
- 102100036011 T-cell surface glycoprotein CD4 Human genes 0.000 description 61
- 239000013598 vector Substances 0.000 description 61
- 239000003981 vehicle Substances 0.000 description 59
- 108020004414 DNA Proteins 0.000 description 55
- 108091033409 CRISPR Proteins 0.000 description 50
- 102100038319 Myosin-6 Human genes 0.000 description 47
- 101710204027 Myosin-6 Proteins 0.000 description 47
- 238000012239 gene modification Methods 0.000 description 40
- 230000005017 genetic modification Effects 0.000 description 40
- 235000013617 genetically modified food Nutrition 0.000 description 40
- 210000002216 heart Anatomy 0.000 description 40
- 238000011282 treatment Methods 0.000 description 39
- 150000007523 nucleic acids Chemical class 0.000 description 37
- 102000004196 processed proteins & peptides Human genes 0.000 description 37
- 241000282414 Homo sapiens Species 0.000 description 33
- 244000005700 microbiome Species 0.000 description 31
- 241000700605 Viruses Species 0.000 description 29
- 108020004707 nucleic acids Proteins 0.000 description 29
- 102000039446 nucleic acids Human genes 0.000 description 29
- -1 ceftamere Chemical compound 0.000 description 26
- 230000002829 reductive effect Effects 0.000 description 26
- 241001515965 unidentified phage Species 0.000 description 26
- 238000004458 analytical method Methods 0.000 description 25
- 102000004190 Enzymes Human genes 0.000 description 24
- 108090000790 Enzymes Proteins 0.000 description 24
- 108010005774 beta-Galactosidase Proteins 0.000 description 24
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 24
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 24
- 229940088598 enzyme Drugs 0.000 description 24
- 239000002953 phosphate buffered saline Substances 0.000 description 24
- 239000013612 plasmid Substances 0.000 description 24
- 239000002245 particle Substances 0.000 description 22
- 201000010099 disease Diseases 0.000 description 20
- 230000002550 fecal effect Effects 0.000 description 19
- 238000000684 flow cytometry Methods 0.000 description 19
- 239000000523 sample Substances 0.000 description 19
- 238000012546 transfer Methods 0.000 description 19
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 18
- 108700028369 Alleles Proteins 0.000 description 18
- 230000003278 mimic effect Effects 0.000 description 18
- 108020005004 Guide RNA Proteins 0.000 description 17
- 239000007788 liquid Substances 0.000 description 17
- 230000027455 binding Effects 0.000 description 16
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Chemical compound NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 16
- 238000002474 experimental method Methods 0.000 description 16
- 230000009257 reactivity Effects 0.000 description 16
- 108010065026 HLA-DQB1 antigen Proteins 0.000 description 15
- 230000000813 microbial effect Effects 0.000 description 15
- 206010012812 Diffuse cutaneous mastocytosis Diseases 0.000 description 14
- 102100036241 HLA class II histocompatibility antigen, DQ beta 1 chain Human genes 0.000 description 14
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 13
- 229940098773 bovine serum albumin Drugs 0.000 description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 13
- 241001465754 Metazoa Species 0.000 description 12
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 12
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 12
- 239000011324 bead Substances 0.000 description 12
- WQZGKKKJIJFFOK-FPRJBGLDSA-N beta-D-galactose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-FPRJBGLDSA-N 0.000 description 12
- 238000010790 dilution Methods 0.000 description 12
- 239000012895 dilution Substances 0.000 description 12
- 230000000694 effects Effects 0.000 description 12
- 230000000968 intestinal effect Effects 0.000 description 12
- 238000012360 testing method Methods 0.000 description 12
- 102100031780 Endonuclease Human genes 0.000 description 11
- 206010061218 Inflammation Diseases 0.000 description 11
- 102100037850 Interferon gamma Human genes 0.000 description 11
- 108010074328 Interferon-gamma Proteins 0.000 description 11
- 238000012217 deletion Methods 0.000 description 11
- 230000002068 genetic effect Effects 0.000 description 11
- 230000004054 inflammatory process Effects 0.000 description 11
- 210000004400 mucous membrane Anatomy 0.000 description 11
- 239000006041 probiotic Substances 0.000 description 11
- 235000018291 probiotics Nutrition 0.000 description 11
- 230000035755 proliferation Effects 0.000 description 11
- 238000012163 sequencing technique Methods 0.000 description 11
- 238000007619 statistical method Methods 0.000 description 11
- 108010008532 Deoxyribonuclease I Proteins 0.000 description 10
- 102000007260 Deoxyribonuclease I Human genes 0.000 description 10
- 206010019280 Heart failures Diseases 0.000 description 10
- 102000013691 Interleukin-17 Human genes 0.000 description 10
- 108050003558 Interleukin-17 Proteins 0.000 description 10
- 108091028043 Nucleic acid sequence Proteins 0.000 description 10
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 description 10
- 230000004913 activation Effects 0.000 description 10
- 150000001413 amino acids Chemical class 0.000 description 10
- 230000037430 deletion Effects 0.000 description 10
- 238000001543 one-way ANOVA Methods 0.000 description 10
- 210000002966 serum Anatomy 0.000 description 10
- 241000736262 Microbiota Species 0.000 description 9
- 235000001014 amino acid Nutrition 0.000 description 9
- 229940024606 amino acid Drugs 0.000 description 9
- 102000005936 beta-Galactosidase Human genes 0.000 description 9
- 208000015181 infectious disease Diseases 0.000 description 9
- 230000004048 modification Effects 0.000 description 9
- 238000012986 modification Methods 0.000 description 9
- 239000007787 solid Substances 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- 241000894007 species Species 0.000 description 9
- 241000606215 Bacteroides vulgatus Species 0.000 description 8
- 238000010354 CRISPR gene editing Methods 0.000 description 8
- 238000002965 ELISA Methods 0.000 description 8
- ULGZDMOVFRHVEP-RWJQBGPGSA-N Erythromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 ULGZDMOVFRHVEP-RWJQBGPGSA-N 0.000 description 8
- 101000738771 Homo sapiens Receptor-type tyrosine-protein phosphatase C Proteins 0.000 description 8
- 102100037422 Receptor-type tyrosine-protein phosphatase C Human genes 0.000 description 8
- 238000000692 Student's t-test Methods 0.000 description 8
- 229940104302 cytosine Drugs 0.000 description 8
- 239000012634 fragment Substances 0.000 description 8
- 238000004519 manufacturing process Methods 0.000 description 8
- 238000002844 melting Methods 0.000 description 8
- 230000008018 melting Effects 0.000 description 8
- 238000012216 screening Methods 0.000 description 8
- 238000011725 BALB/c mouse Methods 0.000 description 7
- 108700004991 Cas12a Proteins 0.000 description 7
- 102210010092 DQB1*03:01 Human genes 0.000 description 7
- 241000588697 Enterobacter cloacae Species 0.000 description 7
- 238000009825 accumulation Methods 0.000 description 7
- 210000001072 colon Anatomy 0.000 description 7
- 229920001577 copolymer Polymers 0.000 description 7
- 235000013305 food Nutrition 0.000 description 7
- 238000000338 in vitro Methods 0.000 description 7
- 230000002147 killing effect Effects 0.000 description 7
- 210000001165 lymph node Anatomy 0.000 description 7
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 7
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 7
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 7
- 239000000843 powder Substances 0.000 description 7
- 238000010186 staining Methods 0.000 description 7
- 239000000725 suspension Substances 0.000 description 7
- 229930024421 Adenine Natural products 0.000 description 6
- 102000055025 Adenosine deaminases Human genes 0.000 description 6
- 241000588724 Escherichia coli Species 0.000 description 6
- 241000606210 Parabacteroides distasonis Species 0.000 description 6
- ISAKRJDGNUQOIC-UHFFFAOYSA-N Uracil Chemical compound O=C1C=CNC(=O)N1 ISAKRJDGNUQOIC-UHFFFAOYSA-N 0.000 description 6
- 229960000643 adenine Drugs 0.000 description 6
- 210000000234 capsid Anatomy 0.000 description 6
- 230000000747 cardiac effect Effects 0.000 description 6
- 239000006071 cream Substances 0.000 description 6
- 230000003111 delayed effect Effects 0.000 description 6
- 230000002939 deleterious effect Effects 0.000 description 6
- 238000002405 diagnostic procedure Methods 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 238000003114 enzyme-linked immunosorbent spot assay Methods 0.000 description 6
- 210000003608 fece Anatomy 0.000 description 6
- 230000006870 function Effects 0.000 description 6
- 102000054766 genetic haplotypes Human genes 0.000 description 6
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 6
- 238000001727 in vivo Methods 0.000 description 6
- 230000003993 interaction Effects 0.000 description 6
- 230000035772 mutation Effects 0.000 description 6
- 239000008188 pellet Substances 0.000 description 6
- 229920001184 polypeptide Polymers 0.000 description 6
- 230000000529 probiotic effect Effects 0.000 description 6
- 238000011002 quantification Methods 0.000 description 6
- 230000010076 replication Effects 0.000 description 6
- 230000004044 response Effects 0.000 description 6
- 239000007921 spray Substances 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 5
- 108020000946 Bacterial DNA Proteins 0.000 description 5
- 108010074051 C-Reactive Protein Proteins 0.000 description 5
- 102100032752 C-reactive protein Human genes 0.000 description 5
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 5
- 102000053602 DNA Human genes 0.000 description 5
- 102210047287 DQB1*03:02 Human genes 0.000 description 5
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 5
- 241000699660 Mus musculus Species 0.000 description 5
- 102000003505 Myosin Human genes 0.000 description 5
- 108060008487 Myosin Proteins 0.000 description 5
- 108091030145 Retron msr RNA Proteins 0.000 description 5
- 229960005305 adenosine Drugs 0.000 description 5
- 239000000443 aerosol Substances 0.000 description 5
- 210000004369 blood Anatomy 0.000 description 5
- 239000008280 blood Substances 0.000 description 5
- 239000002775 capsule Substances 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- 230000000875 corresponding effect Effects 0.000 description 5
- 230000016396 cytokine production Effects 0.000 description 5
- 230000001419 dependent effect Effects 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 239000003814 drug Substances 0.000 description 5
- 239000000975 dye Substances 0.000 description 5
- 230000004927 fusion Effects 0.000 description 5
- 108020001507 fusion proteins Proteins 0.000 description 5
- 102000037865 fusion proteins Human genes 0.000 description 5
- 239000000499 gel Substances 0.000 description 5
- 238000010362 genome editing Methods 0.000 description 5
- 230000012010 growth Effects 0.000 description 5
- 208000019622 heart disease Diseases 0.000 description 5
- 238000011534 incubation Methods 0.000 description 5
- 230000007246 mechanism Effects 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 210000000066 myeloid cell Anatomy 0.000 description 5
- 230000006780 non-homologous end joining Effects 0.000 description 5
- 125000003729 nucleotide group Chemical group 0.000 description 5
- 239000002674 ointment Substances 0.000 description 5
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 description 5
- 229920001223 polyethylene glycol Polymers 0.000 description 5
- 210000000952 spleen Anatomy 0.000 description 5
- 210000004988 splenocyte Anatomy 0.000 description 5
- 235000000346 sugar Nutrition 0.000 description 5
- 229960004673 sulfadoxine Drugs 0.000 description 5
- 239000003826 tablet Substances 0.000 description 5
- 210000001519 tissue Anatomy 0.000 description 5
- 239000001993 wax Substances 0.000 description 5
- 108020004465 16S ribosomal RNA Proteins 0.000 description 4
- 101710169336 5'-deoxyadenosine deaminase Proteins 0.000 description 4
- 108010079649 APOBEC-1 Deaminase Proteins 0.000 description 4
- 102000012758 APOBEC-1 Deaminase Human genes 0.000 description 4
- 206010064539 Autoimmune myocarditis Diseases 0.000 description 4
- 101100491335 Caenorhabditis elegans mat-2 gene Proteins 0.000 description 4
- 108091026890 Coding region Proteins 0.000 description 4
- 102210047284 DQA1*03:01 Human genes 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 108010010803 Gelatin Proteins 0.000 description 4
- AEMRFAOFKBGASW-UHFFFAOYSA-N Glycolic acid Chemical compound OCC(O)=O AEMRFAOFKBGASW-UHFFFAOYSA-N 0.000 description 4
- 241000282412 Homo Species 0.000 description 4
- 229930010555 Inosine Natural products 0.000 description 4
- UGQMRVRMYYASKQ-KQYNXXCUSA-N Inosine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C2=NC=NC(O)=C2N=C1 UGQMRVRMYYASKQ-KQYNXXCUSA-N 0.000 description 4
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N Lactic Acid Natural products CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 4
- 102000043131 MHC class II family Human genes 0.000 description 4
- 108091054438 MHC class II family Proteins 0.000 description 4
- 241001529936 Murinae Species 0.000 description 4
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 4
- 230000026279 RNA modification Effects 0.000 description 4
- 238000011529 RT qPCR Methods 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 229920002472 Starch Polymers 0.000 description 4
- 210000000068 Th17 cell Anatomy 0.000 description 4
- 201000005180 acute myocarditis Diseases 0.000 description 4
- 230000000844 anti-bacterial effect Effects 0.000 description 4
- 230000005875 antibody response Effects 0.000 description 4
- 239000000427 antigen Substances 0.000 description 4
- 210000003719 b-lymphocyte Anatomy 0.000 description 4
- 230000008901 benefit Effects 0.000 description 4
- 230000033228 biological regulation Effects 0.000 description 4
- 238000001574 biopsy Methods 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 239000006285 cell suspension Substances 0.000 description 4
- 239000001913 cellulose Substances 0.000 description 4
- 230000002860 competitive effect Effects 0.000 description 4
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 4
- 235000018417 cysteine Nutrition 0.000 description 4
- 230000034994 death Effects 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 235000014113 dietary fatty acids Nutrition 0.000 description 4
- 208000035475 disorder Diseases 0.000 description 4
- 230000005782 double-strand break Effects 0.000 description 4
- 239000012636 effector Substances 0.000 description 4
- 229960003276 erythromycin Drugs 0.000 description 4
- AEUTYOVWOVBAKS-UWVGGRQHSA-N ethambutol Chemical compound CC[C@@H](CO)NCCN[C@@H](CC)CO AEUTYOVWOVBAKS-UWVGGRQHSA-N 0.000 description 4
- 230000003203 everyday effect Effects 0.000 description 4
- 239000000194 fatty acid Substances 0.000 description 4
- 229930195729 fatty acid Natural products 0.000 description 4
- ODKNJVUHOIMIIZ-RRKCRQDMSA-N floxuridine Chemical compound C1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C(F)=C1 ODKNJVUHOIMIIZ-RRKCRQDMSA-N 0.000 description 4
- 239000008273 gelatin Substances 0.000 description 4
- 229920000159 gelatin Polymers 0.000 description 4
- 229940014259 gelatin Drugs 0.000 description 4
- 235000019322 gelatine Nutrition 0.000 description 4
- 235000011852 gelatine desserts Nutrition 0.000 description 4
- 230000004217 heart function Effects 0.000 description 4
- 210000005003 heart tissue Anatomy 0.000 description 4
- 230000006801 homologous recombination Effects 0.000 description 4
- 238000002744 homologous recombination Methods 0.000 description 4
- 235000010979 hydroxypropyl methyl cellulose Nutrition 0.000 description 4
- 239000001866 hydroxypropyl methyl cellulose Substances 0.000 description 4
- 229920003088 hydroxypropyl methyl cellulose Polymers 0.000 description 4
- UFVKGYZPFZQRLF-UHFFFAOYSA-N hydroxypropyl methyl cellulose Chemical compound OC1C(O)C(OC)OC(CO)C1OC1C(O)C(O)C(OC2C(C(O)C(OC3C(C(O)C(O)C(CO)O3)O)C(CO)O2)O)C(CO)O1 UFVKGYZPFZQRLF-UHFFFAOYSA-N 0.000 description 4
- 210000002865 immune cell Anatomy 0.000 description 4
- 230000008676 import Effects 0.000 description 4
- 230000001939 inductive effect Effects 0.000 description 4
- 229960003786 inosine Drugs 0.000 description 4
- 238000003780 insertion Methods 0.000 description 4
- 230000037431 insertion Effects 0.000 description 4
- 239000006210 lotion Substances 0.000 description 4
- 230000002101 lytic effect Effects 0.000 description 4
- 210000002540 macrophage Anatomy 0.000 description 4
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 239000002773 nucleotide Substances 0.000 description 4
- 244000052769 pathogen Species 0.000 description 4
- 229920000642 polymer Polymers 0.000 description 4
- 239000003755 preservative agent Substances 0.000 description 4
- 230000009467 reduction Effects 0.000 description 4
- 230000002441 reversible effect Effects 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 239000002904 solvent Substances 0.000 description 4
- 235000019698 starch Nutrition 0.000 description 4
- 239000008107 starch Substances 0.000 description 4
- 230000000638 stimulation Effects 0.000 description 4
- 230000004083 survival effect Effects 0.000 description 4
- 201000000596 systemic lupus erythematosus Diseases 0.000 description 4
- RWQNBRDOKXIBIV-UHFFFAOYSA-N thymine Chemical compound CC1=CNC(=O)NC1=O RWQNBRDOKXIBIV-UHFFFAOYSA-N 0.000 description 4
- 238000011830 transgenic mouse model Methods 0.000 description 4
- 238000002054 transplantation Methods 0.000 description 4
- 238000011144 upstream manufacturing Methods 0.000 description 4
- 230000002861 ventricular Effects 0.000 description 4
- 238000005406 washing Methods 0.000 description 4
- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 3
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 3
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 229920002126 Acrylic acid copolymer Polymers 0.000 description 3
- 241000606124 Bacteroides fragilis Species 0.000 description 3
- 241001221145 Bacteroides pyogenes Species 0.000 description 3
- 101100284398 Bos taurus BoLA-DQB gene Proteins 0.000 description 3
- 102000004127 Cytokines Human genes 0.000 description 3
- 108090000695 Cytokines Proteins 0.000 description 3
- 102000000311 Cytosine Deaminase Human genes 0.000 description 3
- 108010080611 Cytosine Deaminase Proteins 0.000 description 3
- 102210047546 DQB1*03:03 Human genes 0.000 description 3
- 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 3
- 241000588914 Enterobacter Species 0.000 description 3
- 108091029865 Exogenous DNA Proteins 0.000 description 3
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 3
- 239000007995 HEPES buffer Substances 0.000 description 3
- 108010062347 HLA-DQ Antigens Proteins 0.000 description 3
- 108010047762 HLA-DQ8 antigen Proteins 0.000 description 3
- 102000004157 Hydrolases Human genes 0.000 description 3
- 108090000604 Hydrolases Proteins 0.000 description 3
- 229920002153 Hydroxypropyl cellulose Polymers 0.000 description 3
- 229940076838 Immune checkpoint inhibitor Drugs 0.000 description 3
- 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 3
- OFOBLEOULBTSOW-UHFFFAOYSA-N Malonic acid Chemical compound OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 3
- 241000124008 Mammalia Species 0.000 description 3
- 241000699666 Mus <mouse, genus> Species 0.000 description 3
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- ZVGNESXIJDCBKN-WUIGKKEISA-N R-Tiacumicin B Natural products O([C@@H]1[C@@H](C)O[C@H]([C@H]([C@H]1O)OC)OCC1=CC=CC[C@H](O)C(C)=C[C@@H]([C@H](C(C)=CC(C)=CC[C@H](OC1=O)[C@@H](C)O)O[C@H]1[C@H]([C@@H](O)[C@H](OC(=O)C(C)C)C(C)(C)O1)O)CC)C(=O)C1=C(O)C(Cl)=C(O)C(Cl)=C1CC ZVGNESXIJDCBKN-WUIGKKEISA-N 0.000 description 3
- 238000010357 RNA editing Methods 0.000 description 3
- 241000700159 Rattus Species 0.000 description 3
- 235000021355 Stearic acid Nutrition 0.000 description 3
- 230000005867 T cell response Effects 0.000 description 3
- 239000004098 Tetracycline Substances 0.000 description 3
- 210000000447 Th1 cell Anatomy 0.000 description 3
- 108091007433 antigens Proteins 0.000 description 3
- 102000036639 antigens Human genes 0.000 description 3
- 238000013459 approach Methods 0.000 description 3
- 230000033590 base-excision repair Effects 0.000 description 3
- 239000000969 carrier Substances 0.000 description 3
- 235000010980 cellulose Nutrition 0.000 description 3
- 229920002678 cellulose Polymers 0.000 description 3
- 229920002301 cellulose acetate Polymers 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 210000004443 dendritic cell Anatomy 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 239000000835 fiber Substances 0.000 description 3
- 229960000628 fidaxomicin Drugs 0.000 description 3
- ZVGNESXIJDCBKN-UUEYKCAUSA-N fidaxomicin Chemical compound O([C@@H]1[C@@H](C)O[C@H]([C@H]([C@H]1O)OC)OCC\1=C/C=C/C[C@H](O)/C(C)=C/[C@@H]([C@H](/C(C)=C/C(/C)=C/C[C@H](OC/1=O)[C@@H](C)O)O[C@H]1[C@H]([C@@H](O)[C@H](OC(=O)C(C)C)C(C)(C)O1)O)CC)C(=O)C1=C(O)C(Cl)=C(O)C(Cl)=C1CC ZVGNESXIJDCBKN-UUEYKCAUSA-N 0.000 description 3
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 3
- 238000009472 formulation Methods 0.000 description 3
- 235000010977 hydroxypropyl cellulose Nutrition 0.000 description 3
- 239000001863 hydroxypropyl cellulose Substances 0.000 description 3
- 239000012274 immune-checkpoint protein inhibitor Substances 0.000 description 3
- 230000006698 induction Effects 0.000 description 3
- 230000024949 interleukin-17 production Effects 0.000 description 3
- 239000008101 lactose Substances 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- 230000004060 metabolic process Effects 0.000 description 3
- 210000004165 myocardium Anatomy 0.000 description 3
- GLDOVTGHNKAZLK-UHFFFAOYSA-N octadecan-1-ol Chemical compound CCCCCCCCCCCCCCCCCCO GLDOVTGHNKAZLK-UHFFFAOYSA-N 0.000 description 3
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 3
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 3
- 239000003921 oil Substances 0.000 description 3
- 235000019198 oils Nutrition 0.000 description 3
- 210000000056 organ Anatomy 0.000 description 3
- 239000003002 pH adjusting agent Substances 0.000 description 3
- 230000001717 pathogenic effect Effects 0.000 description 3
- 235000019271 petrolatum Nutrition 0.000 description 3
- 239000008177 pharmaceutical agent Substances 0.000 description 3
- 239000000546 pharmaceutical excipient Substances 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 230000000750 progressive effect Effects 0.000 description 3
- 239000003380 propellant Substances 0.000 description 3
- 102000005962 receptors Human genes 0.000 description 3
- 108020003175 receptors Proteins 0.000 description 3
- 230000008439 repair process Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 238000004904 shortening Methods 0.000 description 3
- 239000008117 stearic acid Substances 0.000 description 3
- 239000000829 suppository Substances 0.000 description 3
- 239000000375 suspending agent Substances 0.000 description 3
- 230000009885 systemic effect Effects 0.000 description 3
- 230000008685 targeting Effects 0.000 description 3
- 235000019364 tetracycline Nutrition 0.000 description 3
- 150000003522 tetracyclines Chemical class 0.000 description 3
- 238000002560 therapeutic procedure Methods 0.000 description 3
- 230000000699 topical effect Effects 0.000 description 3
- 231100000331 toxic Toxicity 0.000 description 3
- 230000002588 toxic effect Effects 0.000 description 3
- 229940035893 uracil Drugs 0.000 description 3
- 210000002700 urine Anatomy 0.000 description 3
- 230000003612 virological effect Effects 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- IXXFZUPTQVDPPK-ZAWHAJPISA-N (1r,2r,4r,6r,7r,8r,10s,13r,14s)-17-[4-[4-(3-aminophenyl)triazol-1-yl]butyl]-7-[(2s,3r,4s,6r)-4-(dimethylamino)-3-hydroxy-6-methyloxan-2-yl]oxy-13-ethyl-10-fluoro-6-methoxy-2,4,6,8,10,14-hexamethyl-12,15-dioxa-17-azabicyclo[12.3.0]heptadecane-3,9,11,16-tet Chemical compound O([C@@H]1[C@@H](C)C(=O)[C@](C)(F)C(=O)O[C@@H]([C@]2(OC(=O)N(CCCCN3N=NC(=C3)C=3C=C(N)C=CC=3)[C@@H]2[C@@H](C)C(=O)[C@H](C)C[C@@]1(C)OC)C)CC)[C@@H]1O[C@H](C)C[C@H](N(C)C)[C@H]1O IXXFZUPTQVDPPK-ZAWHAJPISA-N 0.000 description 2
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 description 2
- 102000040650 (ribonucleotides)n+m Human genes 0.000 description 2
- GOJUJUVQIVIZAV-UHFFFAOYSA-N 2-amino-4,6-dichloropyrimidine-5-carbaldehyde Chemical group NC1=NC(Cl)=C(C=O)C(Cl)=N1 GOJUJUVQIVIZAV-UHFFFAOYSA-N 0.000 description 2
- RYVNIFSIEDRLSJ-UHFFFAOYSA-N 5-(hydroxymethyl)cytosine Chemical compound NC=1NC(=O)N=CC=1CO RYVNIFSIEDRLSJ-UHFFFAOYSA-N 0.000 description 2
- FFKUHGONCHRHPE-UHFFFAOYSA-N 5-methyl-1h-pyrimidine-2,4-dione;7h-purin-6-amine Chemical compound CC1=CNC(=O)NC1=O.NC1=NC=NC2=C1NC=N2 FFKUHGONCHRHPE-UHFFFAOYSA-N 0.000 description 2
- LRFVTYWOQMYALW-UHFFFAOYSA-N 9H-xanthine Chemical compound O=C1NC(=O)NC2=C1NC=N2 LRFVTYWOQMYALW-UHFFFAOYSA-N 0.000 description 2
- 108091093088 Amplicon Proteins 0.000 description 2
- 208000023275 Autoimmune disease Diseases 0.000 description 2
- 241000186000 Bifidobacterium Species 0.000 description 2
- 102100031151 C-C chemokine receptor type 2 Human genes 0.000 description 2
- 101710149815 C-C chemokine receptor type 2 Proteins 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- 108010051609 Cardiac Myosins Proteins 0.000 description 2
- 102000013602 Cardiac Myosins Human genes 0.000 description 2
- 208000020446 Cardiac disease Diseases 0.000 description 2
- 206010048610 Cardiotoxicity Diseases 0.000 description 2
- 108010078791 Carrier Proteins Proteins 0.000 description 2
- GNWUOVJNSFPWDD-XMZRARIVSA-M Cefoxitin sodium Chemical compound [Na+].N([C@]1(OC)C(N2C(=C(COC(N)=O)CS[C@@H]21)C([O-])=O)=O)C(=O)CC1=CC=CS1 GNWUOVJNSFPWDD-XMZRARIVSA-M 0.000 description 2
- 108010073254 Colicins Proteins 0.000 description 2
- 102000029816 Collagenase Human genes 0.000 description 2
- 108060005980 Collagenase Proteins 0.000 description 2
- 102000005381 Cytidine Deaminase Human genes 0.000 description 2
- 108010031325 Cytidine deaminase Proteins 0.000 description 2
- 101710180243 Cytidine deaminase 1 Proteins 0.000 description 2
- RGHNJXZEOKUKBD-SQOUGZDYSA-N D-gluconic acid Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 description 2
- 102100040263 DNA dC->dU-editing enzyme APOBEC-3A Human genes 0.000 description 2
- 102210047476 DQA1*02:01 Human genes 0.000 description 2
- 239000001856 Ethyl cellulose Substances 0.000 description 2
- ZZSNKZQZMQGXPY-UHFFFAOYSA-N Ethyl cellulose Chemical compound CCOCC1OC(OC)C(OCC)C(OCC)C1OC1C(O)C(O)C(OC)C(CO)O1 ZZSNKZQZMQGXPY-UHFFFAOYSA-N 0.000 description 2
- 241000589599 Francisella tularensis subsp. novicida Species 0.000 description 2
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 description 2
- 101150106478 GPS1 gene Proteins 0.000 description 2
- 102000002464 Galactosidases Human genes 0.000 description 2
- 108010093031 Galactosidases Proteins 0.000 description 2
- 208000034826 Genetic Predisposition to Disease Diseases 0.000 description 2
- 229930182566 Gentamicin Natural products 0.000 description 2
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 2
- 102100036243 HLA class II histocompatibility antigen, DQ alpha 1 chain Human genes 0.000 description 2
- 108010086786 HLA-DQA1 antigen Proteins 0.000 description 2
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 2
- 101000964378 Homo sapiens DNA dC->dU-editing enzyme APOBEC-3A Proteins 0.000 description 2
- 102000037984 Inhibitory immune checkpoint proteins Human genes 0.000 description 2
- 108091008026 Inhibitory immune checkpoint proteins Proteins 0.000 description 2
- 229920001202 Inulin Polymers 0.000 description 2
- 241000186660 Lactobacillus Species 0.000 description 2
- OJMMVQQUTAEWLP-UHFFFAOYSA-N Lincomycin Natural products CN1CC(CCC)CC1C(=O)NC(C(C)O)C1C(O)C(O)C(O)C(SC)O1 OJMMVQQUTAEWLP-UHFFFAOYSA-N 0.000 description 2
- 238000000585 Mann–Whitney U test Methods 0.000 description 2
- MJVAVZPDRWSRRC-UHFFFAOYSA-N Menadione Chemical compound C1=CC=C2C(=O)C(C)=CC(=O)C2=C1 MJVAVZPDRWSRRC-UHFFFAOYSA-N 0.000 description 2
- VVQNEPGJFQJSBK-UHFFFAOYSA-N Methyl methacrylate Chemical compound COC(=O)C(C)=C VVQNEPGJFQJSBK-UHFFFAOYSA-N 0.000 description 2
- 229920000168 Microcrystalline cellulose Polymers 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- 241000283973 Oryctolagus cuniculus Species 0.000 description 2
- 108010058846 Ovalbumin Proteins 0.000 description 2
- 102000004316 Oxidoreductases Human genes 0.000 description 2
- 108090000854 Oxidoreductases Proteins 0.000 description 2
- 238000012408 PCR amplification Methods 0.000 description 2
- 241000160321 Parabacteroides Species 0.000 description 2
- 241001494479 Pecora Species 0.000 description 2
- 239000004264 Petrolatum Substances 0.000 description 2
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 2
- 239000002202 Polyethylene glycol Substances 0.000 description 2
- 108010025955 Pyocins Proteins 0.000 description 2
- 239000012980 RPMI-1640 medium Substances 0.000 description 2
- 108091028664 Ribonucleotide Proteins 0.000 description 2
- 239000006146 Roswell Park Memorial Institute medium Substances 0.000 description 2
- 241001515849 Satellite Viruses Species 0.000 description 2
- 241000191967 Staphylococcus aureus Species 0.000 description 2
- 241000193996 Streptococcus pyogenes Species 0.000 description 2
- 101710172711 Structural protein Proteins 0.000 description 2
- NHUHCSRWZMLRLA-UHFFFAOYSA-N Sulfisoxazole Chemical compound CC1=NOC(NS(=O)(=O)C=2C=CC(N)=CC=2)=C1C NHUHCSRWZMLRLA-UHFFFAOYSA-N 0.000 description 2
- 108091028113 Trans-activating crRNA Proteins 0.000 description 2
- 102100022356 Tyrosine-protein kinase Mer Human genes 0.000 description 2
- 101710117021 Tyrosine-protein phosphatase YopH Proteins 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- WNLRTRBMVRJNCN-UHFFFAOYSA-N adipic acid Chemical compound OC(=O)CCCCC(O)=O WNLRTRBMVRJNCN-UHFFFAOYSA-N 0.000 description 2
- 238000013019 agitation Methods 0.000 description 2
- 150000001298 alcohols Chemical class 0.000 description 2
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 description 2
- 229920003144 amino alkyl methacrylate copolymer Polymers 0.000 description 2
- 229940126575 aminoglycoside Drugs 0.000 description 2
- 238000002583 angiography Methods 0.000 description 2
- 230000007503 antigenic stimulation Effects 0.000 description 2
- 230000001363 autoimmune Effects 0.000 description 2
- 230000005784 autoimmunity Effects 0.000 description 2
- 210000003578 bacterial chromosome Anatomy 0.000 description 2
- 235000013871 bee wax Nutrition 0.000 description 2
- 239000012166 beeswax Substances 0.000 description 2
- WPYMKLBDIGXBTP-UHFFFAOYSA-N benzoic acid Chemical compound OC(=O)C1=CC=CC=C1 WPYMKLBDIGXBTP-UHFFFAOYSA-N 0.000 description 2
- 239000011230 binding agent Substances 0.000 description 2
- 238000003766 bioinformatics method Methods 0.000 description 2
- 210000004556 brain Anatomy 0.000 description 2
- 108010018804 c-Mer Tyrosine Kinase Proteins 0.000 description 2
- 239000001506 calcium phosphate Substances 0.000 description 2
- 229910000389 calcium phosphate Inorganic materials 0.000 description 2
- 235000011010 calcium phosphates Nutrition 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 description 2
- 238000013184 cardiac magnetic resonance imaging Methods 0.000 description 2
- 231100000259 cardiotoxicity Toxicity 0.000 description 2
- 239000004203 carnauba wax Substances 0.000 description 2
- 235000013869 carnauba wax Nutrition 0.000 description 2
- 239000004359 castor oil Substances 0.000 description 2
- 235000019438 castor oil Nutrition 0.000 description 2
- 229960005495 cefotetan Drugs 0.000 description 2
- SRZNHPXWXCNNDU-RHBCBLIFSA-N cefotetan Chemical compound N([C@]1(OC)C(N2C(=C(CSC=3N(N=NN=3)C)CS[C@@H]21)C(O)=O)=O)C(=O)C1SC(=C(C(N)=O)C(O)=O)S1 SRZNHPXWXCNNDU-RHBCBLIFSA-N 0.000 description 2
- 229960002682 cefoxitin Drugs 0.000 description 2
- 230000030833 cell death Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000001684 chronic effect Effects 0.000 description 2
- MYSWGUAQZAJSOK-UHFFFAOYSA-N ciprofloxacin Chemical compound C12=CC(N3CCNCC3)=C(F)C=C2C(=O)C(C(=O)O)=CN1C1CC1 MYSWGUAQZAJSOK-UHFFFAOYSA-N 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 229960002424 collagenase Drugs 0.000 description 2
- 239000003086 colorant Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000021615 conjugation Effects 0.000 description 2
- 230000009089 cytolysis Effects 0.000 description 2
- 230000002950 deficient Effects 0.000 description 2
- 239000005547 deoxyribonucleotide Substances 0.000 description 2
- 125000002637 deoxyribonucleotide group Chemical group 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- POULHZVOKOAJMA-UHFFFAOYSA-N dodecanoic acid ester group Chemical group C(CCCCCCCCCCC)(=O)O POULHZVOKOAJMA-UHFFFAOYSA-N 0.000 description 2
- 238000010410 dusting Methods 0.000 description 2
- 239000003995 emulsifying agent Substances 0.000 description 2
- 235000019325 ethyl cellulose Nutrition 0.000 description 2
- 229920001249 ethyl cellulose Polymers 0.000 description 2
- 230000007717 exclusion Effects 0.000 description 2
- 239000003925 fat Substances 0.000 description 2
- 235000019197 fats Nutrition 0.000 description 2
- 150000004665 fatty acids Chemical class 0.000 description 2
- 239000000796 flavoring agent Substances 0.000 description 2
- 235000013355 food flavoring agent Nutrition 0.000 description 2
- 235000003599 food sweetener Nutrition 0.000 description 2
- 229960002518 gentamicin Drugs 0.000 description 2
- ZEMPKEQAKRGZGQ-XOQCFJPHSA-N glycerol triricinoleate Natural products CCCCCC[C@@H](O)CC=CCCCCCCCC(=O)OC[C@@H](COC(=O)CCCCCCCC=CC[C@@H](O)CCCCCC)OC(=O)CCCCCCCC=CC[C@H](O)CCCCCC ZEMPKEQAKRGZGQ-XOQCFJPHSA-N 0.000 description 2
- 244000005709 gut microbiome Species 0.000 description 2
- 229940025294 hemin Drugs 0.000 description 2
- BTIJJDXEELBZFS-QDUVMHSLSA-K hemin Chemical compound CC1=C(CCC(O)=O)C(C=C2C(CCC(O)=O)=C(C)\C(N2[Fe](Cl)N23)=C\4)=N\C1=C/C2=C(C)C(C=C)=C3\C=C/1C(C)=C(C=C)C/4=N\1 BTIJJDXEELBZFS-QDUVMHSLSA-K 0.000 description 2
- BXWNKGSJHAJOGX-UHFFFAOYSA-N hexadecan-1-ol Chemical compound CCCCCCCCCCCCCCCCO BXWNKGSJHAJOGX-UHFFFAOYSA-N 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 239000008172 hydrogenated vegetable oil Substances 0.000 description 2
- 229920001477 hydrophilic polymer Polymers 0.000 description 2
- 230000002209 hydrophobic effect Effects 0.000 description 2
- 229920001600 hydrophobic polymer Polymers 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- 230000001771 impaired effect Effects 0.000 description 2
- 230000001976 improved effect Effects 0.000 description 2
- 210000004969 inflammatory cell Anatomy 0.000 description 2
- 210000002074 inflammatory monocyte Anatomy 0.000 description 2
- 238000001802 infusion Methods 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 244000000056 intracellular parasite Species 0.000 description 2
- JYJIGFIDKWBXDU-MNNPPOADSA-N inulin Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)OC[C@]1(OC[C@]2(OC[C@]3(OC[C@]4(OC[C@]5(OC[C@]6(OC[C@]7(OC[C@]8(OC[C@]9(OC[C@]%10(OC[C@]%11(OC[C@]%12(OC[C@]%13(OC[C@]%14(OC[C@]%15(OC[C@]%16(OC[C@]%17(OC[C@]%18(OC[C@]%19(OC[C@]%20(OC[C@]%21(OC[C@]%22(OC[C@]%23(OC[C@]%24(OC[C@]%25(OC[C@]%26(OC[C@]%27(OC[C@]%28(OC[C@]%29(OC[C@]%30(OC[C@]%31(OC[C@]%32(OC[C@]%33(OC[C@]%34(OC[C@]%35(OC[C@]%36(O[C@@H]%37[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O%37)O)[C@H]([C@H](O)[C@@H](CO)O%36)O)[C@H]([C@H](O)[C@@H](CO)O%35)O)[C@H]([C@H](O)[C@@H](CO)O%34)O)[C@H]([C@H](O)[C@@H](CO)O%33)O)[C@H]([C@H](O)[C@@H](CO)O%32)O)[C@H]([C@H](O)[C@@H](CO)O%31)O)[C@H]([C@H](O)[C@@H](CO)O%30)O)[C@H]([C@H](O)[C@@H](CO)O%29)O)[C@H]([C@H](O)[C@@H](CO)O%28)O)[C@H]([C@H](O)[C@@H](CO)O%27)O)[C@H]([C@H](O)[C@@H](CO)O%26)O)[C@H]([C@H](O)[C@@H](CO)O%25)O)[C@H]([C@H](O)[C@@H](CO)O%24)O)[C@H]([C@H](O)[C@@H](CO)O%23)O)[C@H]([C@H](O)[C@@H](CO)O%22)O)[C@H]([C@H](O)[C@@H](CO)O%21)O)[C@H]([C@H](O)[C@@H](CO)O%20)O)[C@H]([C@H](O)[C@@H](CO)O%19)O)[C@H]([C@H](O)[C@@H](CO)O%18)O)[C@H]([C@H](O)[C@@H](CO)O%17)O)[C@H]([C@H](O)[C@@H](CO)O%16)O)[C@H]([C@H](O)[C@@H](CO)O%15)O)[C@H]([C@H](O)[C@@H](CO)O%14)O)[C@H]([C@H](O)[C@@H](CO)O%13)O)[C@H]([C@H](O)[C@@H](CO)O%12)O)[C@H]([C@H](O)[C@@H](CO)O%11)O)[C@H]([C@H](O)[C@@H](CO)O%10)O)[C@H]([C@H](O)[C@@H](CO)O9)O)[C@H]([C@H](O)[C@@H](CO)O8)O)[C@H]([C@H](O)[C@@H](CO)O7)O)[C@H]([C@H](O)[C@@H](CO)O6)O)[C@H]([C@H](O)[C@@H](CO)O5)O)[C@H]([C@H](O)[C@@H](CO)O4)O)[C@H]([C@H](O)[C@@H](CO)O3)O)[C@H]([C@H](O)[C@@H](CO)O2)O)[C@@H](O)[C@H](O)[C@@H](CO)O1 JYJIGFIDKWBXDU-MNNPPOADSA-N 0.000 description 2
- 229940029339 inulin Drugs 0.000 description 2
- DRAVOWXCEBXPTN-UHFFFAOYSA-N isoguanine Chemical compound NC1=NC(=O)NC2=C1NC=N2 DRAVOWXCEBXPTN-UHFFFAOYSA-N 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 230000003902 lesion Effects 0.000 description 2
- 229960005287 lincomycin Drugs 0.000 description 2
- OJMMVQQUTAEWLP-KIDUDLJLSA-N lincomycin Chemical compound CN1C[C@H](CCC)C[C@H]1C(=O)N[C@H]([C@@H](C)O)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 OJMMVQQUTAEWLP-KIDUDLJLSA-N 0.000 description 2
- 229940041028 lincosamides Drugs 0.000 description 2
- 239000002502 liposome Substances 0.000 description 2
- 229960001977 loracarbef Drugs 0.000 description 2
- JAPHQRWPEGVNBT-UTUOFQBUSA-N loracarbef Chemical compound C1([C@H](C(=O)N[C@@H]2C(N3C(=C(Cl)CC[C@@H]32)C([O-])=O)=O)[NH3+])=CC=CC=C1 JAPHQRWPEGVNBT-UTUOFQBUSA-N 0.000 description 2
- 239000000314 lubricant Substances 0.000 description 2
- 230000001320 lysogenic effect Effects 0.000 description 2
- 235000019359 magnesium stearate Nutrition 0.000 description 2
- 239000001630 malic acid Substances 0.000 description 2
- 235000011090 malic acid Nutrition 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 210000005015 mediastinal lymph node Anatomy 0.000 description 2
- 210000004379 membrane Anatomy 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 229940117841 methacrylic acid copolymer Drugs 0.000 description 2
- 229920003145 methacrylic acid copolymer Polymers 0.000 description 2
- 229920000609 methyl cellulose Polymers 0.000 description 2
- 235000010981 methylcellulose Nutrition 0.000 description 2
- 239000001923 methylcellulose Substances 0.000 description 2
- 229940016286 microcrystalline cellulose Drugs 0.000 description 2
- 235000019813 microcrystalline cellulose Nutrition 0.000 description 2
- 239000008108 microcrystalline cellulose Substances 0.000 description 2
- 238000000386 microscopy Methods 0.000 description 2
- 239000002480 mineral oil Substances 0.000 description 2
- 235000010446 mineral oil Nutrition 0.000 description 2
- 229960000210 nalidixic acid Drugs 0.000 description 2
- MHWLWQUZZRMNGJ-UHFFFAOYSA-N nalidixic acid Chemical compound C1=C(C)N=C2N(CC)C=C(C(O)=O)C(=O)C2=C1 MHWLWQUZZRMNGJ-UHFFFAOYSA-N 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 230000003287 optical effect Effects 0.000 description 2
- 229940092253 ovalbumin Drugs 0.000 description 2
- 239000012188 paraffin wax Substances 0.000 description 2
- 238000007911 parenteral administration Methods 0.000 description 2
- 230000007170 pathology Effects 0.000 description 2
- 239000013610 patient sample Substances 0.000 description 2
- 239000001814 pectin Substances 0.000 description 2
- 235000010987 pectin Nutrition 0.000 description 2
- 229920001277 pectin Polymers 0.000 description 2
- 229940056360 penicillin g Drugs 0.000 description 2
- 229940066842 petrolatum Drugs 0.000 description 2
- 108700010839 phage proteins Proteins 0.000 description 2
- 239000000825 pharmaceutical preparation Substances 0.000 description 2
- PHEDXBVPIONUQT-RGYGYFBISA-N phorbol 13-acetate 12-myristate Chemical compound C([C@]1(O)C(=O)C(C)=C[C@H]1[C@@]1(O)[C@H](C)[C@H]2OC(=O)CCCCCCCCCCCCC)C(CO)=C[C@H]1[C@H]1[C@]2(OC(C)=O)C1(C)C PHEDXBVPIONUQT-RGYGYFBISA-N 0.000 description 2
- 229920003229 poly(methyl methacrylate) Polymers 0.000 description 2
- 229920000193 polymethacrylate Polymers 0.000 description 2
- 239000004926 polymethyl methacrylate Substances 0.000 description 2
- 229920002689 polyvinyl acetate Polymers 0.000 description 2
- 229920002451 polyvinyl alcohol Polymers 0.000 description 2
- 235000019422 polyvinyl alcohol Nutrition 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 235000013406 prebiotics Nutrition 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 230000006798 recombination Effects 0.000 description 2
- 238000005215 recombination Methods 0.000 description 2
- 238000012552 review Methods 0.000 description 2
- 229960001225 rifampicin Drugs 0.000 description 2
- JQXXHWHPUNPDRT-WLSIYKJHSA-N rifampicin Chemical compound O([C@](C1=O)(C)O/C=C/[C@@H]([C@H]([C@@H](OC(C)=O)[C@H](C)[C@H](O)[C@H](C)[C@@H](O)[C@@H](C)\C=C\C=C(C)/C(=O)NC=2C(O)=C3C([O-])=C4C)C)OC)C4=C1C3=C(O)C=2\C=N\N1CC[NH+](C)CC1 JQXXHWHPUNPDRT-WLSIYKJHSA-N 0.000 description 2
- 229960003040 rifaximin Drugs 0.000 description 2
- NZCRJKRKKOLAOJ-XRCRFVBUSA-N rifaximin Chemical compound OC1=C(C(O)=C2C)C3=C4N=C5C=C(C)C=CN5C4=C1NC(=O)\C(C)=C/C=C/[C@H](C)[C@H](O)[C@@H](C)[C@@H](O)[C@@H](C)[C@H](OC(C)=O)[C@H](C)[C@@H](OC)\C=C\O[C@@]1(C)OC2=C3C1=O NZCRJKRKKOLAOJ-XRCRFVBUSA-N 0.000 description 2
- YGSDEFSMJLZEOE-UHFFFAOYSA-N salicylic acid Chemical compound OC(=O)C1=CC=CC=C1O YGSDEFSMJLZEOE-UHFFFAOYSA-N 0.000 description 2
- CXMXRPHRNRROMY-UHFFFAOYSA-N sebacic acid Chemical compound OC(=O)CCCCCCCCC(O)=O CXMXRPHRNRROMY-UHFFFAOYSA-N 0.000 description 2
- 239000008247 solid mixture Substances 0.000 description 2
- 229950008588 solithromycin Drugs 0.000 description 2
- 210000004989 spleen cell Anatomy 0.000 description 2
- 239000008174 sterile solution Substances 0.000 description 2
- 150000008163 sugars Chemical class 0.000 description 2
- 229960000654 sulfafurazole Drugs 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 239000003765 sweetening agent Substances 0.000 description 2
- 229960003250 telithromycin Drugs 0.000 description 2
- LJVAJPDWBABPEJ-PNUFFHFMSA-N telithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)[C@@H](C)C(=O)O[C@@H]([C@]2(OC(=O)N(CCCCN3C=C(N=C3)C=3C=NC=CC=3)[C@@H]2[C@@H](C)C(=O)[C@H](C)C[C@@]1(C)OC)C)CC)[C@@H]1O[C@H](C)C[C@H](N(C)C)[C@H]1O LJVAJPDWBABPEJ-PNUFFHFMSA-N 0.000 description 2
- 229960002180 tetracycline Drugs 0.000 description 2
- 229930101283 tetracycline Natural products 0.000 description 2
- 230000001225 therapeutic effect Effects 0.000 description 2
- 239000002562 thickening agent Substances 0.000 description 2
- 229940113082 thymine Drugs 0.000 description 2
- 108700012359 toxins Proteins 0.000 description 2
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 2
- 230000009466 transformation Effects 0.000 description 2
- 230000009261 transgenic effect Effects 0.000 description 2
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 2
- 230000010415 tropism Effects 0.000 description 2
- 230000035899 viability Effects 0.000 description 2
- 108700026220 vif Genes Proteins 0.000 description 2
- LUEWUZLMQUOBSB-FSKGGBMCSA-N (2s,3s,4s,5s,6r)-2-[(2r,3s,4r,5r,6s)-6-[(2r,3s,4r,5s,6s)-4,5-dihydroxy-2-(hydroxymethyl)-6-[(2r,4r,5s,6r)-4,5,6-trihydroxy-2-(hydroxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-4,5-dihydroxy-2-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol 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]2[C@H](O[C@@H](OC3[C@H](O[C@@H](O)[C@@H](O)[C@H]3O)CO)[C@@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O LUEWUZLMQUOBSB-FSKGGBMCSA-N 0.000 description 1
- HBJOXQRURQPDEX-MHXMMLMNSA-N (2s,4r)-n-[(1s,2s)-2-chloro-1-[(2r,3r,4s,5r,6r)-3,4,5-trihydroxy-6-methylsulfanyloxan-2-yl]propyl]-4-ethylpiperidine-2-carboxamide Chemical compound C1[C@H](CC)CCN[C@@H]1C(=O)N[C@H]([C@H](C)Cl)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 HBJOXQRURQPDEX-MHXMMLMNSA-N 0.000 description 1
- VCOPTHOUUNAYKQ-WBTCAYNUSA-N (3s)-3,6-diamino-n-[[(2s,5s,8e,11s,15s)-15-amino-11-[(6r)-2-amino-1,4,5,6-tetrahydropyrimidin-6-yl]-8-[(carbamoylamino)methylidene]-2-(hydroxymethyl)-3,6,9,12,16-pentaoxo-1,4,7,10,13-pentazacyclohexadec-5-yl]methyl]hexanamide;(3s)-3,6-diamino-n-[[(2s,5s,8 Chemical compound N1C(=O)\C(=C/NC(N)=O)NC(=O)[C@H](CNC(=O)C[C@@H](N)CCCN)NC(=O)[C@H](C)NC(=O)[C@@H](N)CNC(=O)[C@@H]1[C@@H]1NC(N)=NCC1.N1C(=O)\C(=C/NC(N)=O)NC(=O)[C@H](CNC(=O)C[C@@H](N)CCCN)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CNC(=O)[C@@H]1[C@@H]1NC(N)=NCC1 VCOPTHOUUNAYKQ-WBTCAYNUSA-N 0.000 description 1
- SGKRLCUYIXIAHR-AKNGSSGZSA-N (4s,4ar,5s,5ar,6r,12ar)-4-(dimethylamino)-1,5,10,11,12a-pentahydroxy-6-methyl-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1=CC=C2[C@H](C)[C@@H]([C@H](O)[C@@H]3[C@](C(O)=C(C(N)=O)C(=O)[C@H]3N(C)C)(O)C3=O)C3=C(O)C2=C1O SGKRLCUYIXIAHR-AKNGSSGZSA-N 0.000 description 1
- FFTVPQUHLQBXQZ-KVUCHLLUSA-N (4s,4as,5ar,12ar)-4,7-bis(dimethylamino)-1,10,11,12a-tetrahydroxy-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1C2=C(N(C)C)C=CC(O)=C2C(O)=C2[C@@H]1C[C@H]1[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]1(O)C2=O FFTVPQUHLQBXQZ-KVUCHLLUSA-N 0.000 description 1
- SOVUOXKZCCAWOJ-HJYUBDRYSA-N (4s,4as,5ar,12ar)-9-[[2-(tert-butylamino)acetyl]amino]-4,7-bis(dimethylamino)-1,10,11,12a-tetrahydroxy-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1C2=C(N(C)C)C=C(NC(=O)CNC(C)(C)C)C(O)=C2C(O)=C2[C@@H]1C[C@H]1[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]1(O)C2=O SOVUOXKZCCAWOJ-HJYUBDRYSA-N 0.000 description 1
- GUXHBMASAHGULD-SEYHBJAFSA-N (4s,4as,5as,6s,12ar)-7-chloro-4-(dimethylamino)-1,6,10,11,12a-pentahydroxy-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1([C@H]2O)=C(Cl)C=CC(O)=C1C(O)=C1[C@@H]2C[C@H]2[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]2(O)C1=O GUXHBMASAHGULD-SEYHBJAFSA-N 0.000 description 1
- HBUJYEUPIIJJOS-PBHICJAKSA-N (5r)-3-[4-[1-[(2s)-2,3-dihydroxypropanoyl]-3,6-dihydro-2h-pyridin-4-yl]-3,5-difluorophenyl]-5-(1,2-oxazol-3-yloxymethyl)-1,3-oxazolidin-2-one Chemical compound C1N(C(=O)[C@@H](O)CO)CCC(C=2C(=CC(=CC=2F)N2C(O[C@@H](COC3=NOC=C3)C2)=O)F)=C1 HBUJYEUPIIJJOS-PBHICJAKSA-N 0.000 description 1
- MMQXRUYUBYWTDP-KEIGCJKLSA-N (5s,6r,7r)-3-(acetyloxymethyl)-7-[[(2z)-2-(2-amino-1,3-thiazol-4-yl)-2-methoxyiminoacetyl]amino]-5,8-dioxo-5$l^{4}-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound N([C@@H]1C(N2[C@@H]1[S@](CC(COC(C)=O)=C2C(O)=O)=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 MMQXRUYUBYWTDP-KEIGCJKLSA-N 0.000 description 1
- FMZXNVLFJHCSAF-DNVCBOLYSA-N (6R,7R)-3-[(4-carbamoyl-1-pyridin-1-iumyl)methyl]-8-oxo-7-[(1-oxo-2-thiophen-2-ylethyl)amino]-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate Chemical compound C1=CC(C(=O)N)=CC=[N+]1CC1=C(C([O-])=O)N2C(=O)[C@@H](NC(=O)CC=3SC=CC=3)[C@H]2SC1 FMZXNVLFJHCSAF-DNVCBOLYSA-N 0.000 description 1
- YBHZVPYSEUQIII-DYJQDLSISA-N (6r,7r)-3-(acetyloxymethyl)-7-[[(2z)-2-(furan-2-yl)-2-methoxyiminoacetyl]amino]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound N([C@@H]1C(N2C(=C(COC(C)=O)CS[C@@H]21)C(O)=O)=O)C(=O)\C(=N/OC)C1=CC=CO1 YBHZVPYSEUQIII-DYJQDLSISA-N 0.000 description 1
- OCLRGULJISNUQS-OXQOHEQNSA-N (6r,7r)-3-(acetyloxymethyl)-7-[[3-(2-chlorophenyl)-5-methyl-1,2-oxazole-4-carbonyl]amino]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC(=O)C)C(O)=O)C(=O)C1=C(C)ON=C1C1=CC=CC=C1Cl OCLRGULJISNUQS-OXQOHEQNSA-N 0.000 description 1
- LHNIIDJCEODSHA-OQRUQETBSA-N (6r,7r)-3-[(e)-2-(2,4-dinitrophenyl)ethenyl]-8-oxo-7-[(2-thiophen-2-ylacetyl)amino]-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound N([C@H]1[C@H]2SCC(=C(N2C1=O)C(=O)O)\C=C\C=1C(=CC(=CC=1)[N+]([O-])=O)[N+]([O-])=O)C(=O)CC1=CC=CS1 LHNIIDJCEODSHA-OQRUQETBSA-N 0.000 description 1
- QFTZCQVZVRVDTD-DNVCBOLYSA-N (6r,7r)-3-methyl-8-oxo-7-[[2-[4-(1,4,5,6-tetrahydropyrimidin-2-yl)phenyl]acetyl]amino]-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)C)C(O)=O)C(=O)CC(C=C1)=CC=C1C1=NCCCN1 QFTZCQVZVRVDTD-DNVCBOLYSA-N 0.000 description 1
- XSPUSVIQHBDITA-KXDGEKGBSA-N (6r,7r)-7-[[(2e)-2-(2-amino-1,3-thiazol-4-yl)-2-methoxyiminoacetyl]amino]-3-[(5-methyltetrazol-2-yl)methyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)/C(=N/OC)C=2N=C(N)SC=2)CC=1CN1N=NC(C)=N1 XSPUSVIQHBDITA-KXDGEKGBSA-N 0.000 description 1
- RULITNAIJFZYLO-UEKVPHQBSA-N (6r,7r)-7-[[(2r)-2-amino-2-(3-chloro-4-hydroxyphenyl)acetyl]amino]-3-methyl-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)C)C(O)=O)=CC=C(O)C(Cl)=C1 RULITNAIJFZYLO-UEKVPHQBSA-N 0.000 description 1
- SBUCDZYLTRYMFG-PBFPGSCMSA-N (6r,7r)-7-[[(2r)-2-amino-2-(4-hydroxyphenyl)acetyl]amino]-3-[(5-methyl-1,3,4-thiadiazol-2-yl)sulfanylmethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound S1C(C)=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)[C@H](N)C=3C=CC(O)=CC=3)[C@H]2SC1 SBUCDZYLTRYMFG-PBFPGSCMSA-N 0.000 description 1
- WDLWHQDACQUCJR-ZAMMOSSLSA-N (6r,7r)-7-[[(2r)-2-azaniumyl-2-(4-hydroxyphenyl)acetyl]amino]-8-oxo-3-[(e)-prop-1-enyl]-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)/C=C/C)C(O)=O)=CC=C(O)C=C1 WDLWHQDACQUCJR-ZAMMOSSLSA-N 0.000 description 1
- VDFFPBOAOLQAJV-SUYBPPKGSA-N (6r,7r)-7-[[(2r)-2-hydroxy-2-phenylacetyl]amino]-3-[(5-methyl-1,3,4-thiadiazol-2-yl)sulfanylmethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound S1C(C)=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)[C@H](O)C=3C=CC=CC=3)[C@H]2SC1 VDFFPBOAOLQAJV-SUYBPPKGSA-N 0.000 description 1
- YWKJNRNSJKEFMK-PQFQYKRASA-N (6r,7r)-7-[[(2z)-2-(2-amino-1,3-thiazol-4-yl)-2-methoxyiminoacetyl]amino]-8-oxo-3-(5,6,7,8-tetrahydroquinolin-1-ium-1-ylmethyl)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate Chemical compound N([C@@H]1C(N2C(=C(C[N+]=3C=4CCCCC=4C=CC=3)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 YWKJNRNSJKEFMK-PQFQYKRASA-N 0.000 description 1
- CUCFRCCRYQDMNM-PXIRVSTKSA-N (6r,7r)-7-[[(2z)-2-(2-amino-1,3-thiazol-5-yl)-2-(2-oxopyrrolidin-3-yl)oxyiminoacetyl]amino]-8-oxo-3-(pyridin-1-ium-1-ylmethyl)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate Chemical compound S1C(N)=NC=C1C(\C(=O)N[C@@H]1C(N2C(=C(C[N+]=3C=CC=CC=3)CS[C@@H]21)C([O-])=O)=O)=N\OC1C(=O)NCC1 CUCFRCCRYQDMNM-PXIRVSTKSA-N 0.000 description 1
- JXHWKLWIYBATLL-GMIKGCRTSA-N (6r,7r)-7-[[2-[(z)-2-cyanoethenyl]sulfanylacetyl]amino]-3-[(1-methyltetrazol-5-yl)sulfanylmethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound CN1N=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CS\C=C/C#N)[C@H]2SC1 JXHWKLWIYBATLL-GMIKGCRTSA-N 0.000 description 1
- UQWYUAURRDNBKR-BXUZGUMPSA-N (6r,7r)-8-oxo-7-[(2-thiophen-2-ylacetyl)amino]-3-(1h-1,2,4-triazol-5-ylsulfanylmethyl)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid Chemical compound O=C([C@@H](NC(=O)CC=1SC=CC=1)[C@H]1SC2)N1C(C(=O)O)=C2CSC1=NC=NN1 UQWYUAURRDNBKR-BXUZGUMPSA-N 0.000 description 1
- MINDHVHHQZYEEK-UHFFFAOYSA-N (E)-(2S,3R,4R,5S)-5-[(2S,3S,4S,5S)-2,3-epoxy-5-hydroxy-4-methylhexyl]tetrahydro-3,4-dihydroxy-(beta)-methyl-2H-pyran-2-crotonic acid ester with 9-hydroxynonanoic acid Natural products CC(O)C(C)C1OC1CC1C(O)C(O)C(CC(C)=CC(=O)OCCCCCCCCC(O)=O)OC1 MINDHVHHQZYEEK-UHFFFAOYSA-N 0.000 description 1
- RXZBMPWDPOLZGW-XMRMVWPWSA-N (E)-roxithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=N/OCOCCOC)/[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 RXZBMPWDPOLZGW-XMRMVWPWSA-N 0.000 description 1
- QKDHBVNJCZBTMR-LLVKDONJSA-N (R)-temafloxacin Chemical compound C1CN[C@H](C)CN1C(C(=C1)F)=CC2=C1C(=O)C(C(O)=O)=CN2C1=CC=C(F)C=C1F QKDHBVNJCZBTMR-LLVKDONJSA-N 0.000 description 1
- XUBOMFCQGDBHNK-JTQLQIEISA-N (S)-gatifloxacin Chemical compound FC1=CC(C(C(C(O)=O)=CN2C3CC3)=O)=C2C(OC)=C1N1CCN[C@@H](C)C1 XUBOMFCQGDBHNK-JTQLQIEISA-N 0.000 description 1
- YFMFNYKEUDLDTL-UHFFFAOYSA-N 1,1,1,2,3,3,3-heptafluoropropane Chemical compound FC(F)(F)C(F)C(F)(F)F YFMFNYKEUDLDTL-UHFFFAOYSA-N 0.000 description 1
- LVGUZGTVOIAKKC-UHFFFAOYSA-N 1,1,1,2-tetrafluoroethane Chemical compound FCC(F)(F)F LVGUZGTVOIAKKC-UHFFFAOYSA-N 0.000 description 1
- ZORQXIQZAOLNGE-UHFFFAOYSA-N 1,1-difluorocyclohexane Chemical compound FC1(F)CCCCC1 ZORQXIQZAOLNGE-UHFFFAOYSA-N 0.000 description 1
- DDMOUSALMHHKOS-UHFFFAOYSA-N 1,2-dichloro-1,1,2,2-tetrafluoroethane Chemical compound FC(F)(Cl)C(F)(F)Cl DDMOUSALMHHKOS-UHFFFAOYSA-N 0.000 description 1
- GEYOCULIXLDCMW-UHFFFAOYSA-N 1,2-phenylenediamine Chemical compound NC1=CC=CC=C1N GEYOCULIXLDCMW-UHFFFAOYSA-N 0.000 description 1
- OKMWKBLSFKFYGZ-UHFFFAOYSA-N 1-behenoylglycerol Chemical compound CCCCCCCCCCCCCCCCCCCCCC(=O)OCC(O)CO OKMWKBLSFKFYGZ-UHFFFAOYSA-N 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N 2,3,4,5-tetrahydroxypentanal Chemical compound OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- WCOXQTXVACYMLM-UHFFFAOYSA-N 2,3-bis(12-hydroxyoctadecanoyloxy)propyl 12-hydroxyoctadecanoate Chemical compound CCCCCCC(O)CCCCCCCCCCC(=O)OCC(OC(=O)CCCCCCCCCCC(O)CCCCCC)COC(=O)CCCCCCCCCCC(O)CCCCCC WCOXQTXVACYMLM-UHFFFAOYSA-N 0.000 description 1
- SMZOUWXMTYCWNB-UHFFFAOYSA-N 2-(2-methoxy-5-methylphenyl)ethanamine Chemical compound COC1=CC=C(C)C=C1CCN SMZOUWXMTYCWNB-UHFFFAOYSA-N 0.000 description 1
- IZXIZTKNFFYFOF-UHFFFAOYSA-N 2-Oxazolidone Chemical class O=C1NCCO1 IZXIZTKNFFYFOF-UHFFFAOYSA-N 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N 2-Propenoic acid Natural products OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 1
- ACTOXUHEUCPTEW-BWHGAVFKSA-N 2-[(4r,5s,6s,7r,9r,10r,11e,13e,16r)-6-[(2s,3r,4r,5s,6r)-5-[(2s,4r,5s,6s)-4,5-dihydroxy-4,6-dimethyloxan-2-yl]oxy-4-(dimethylamino)-3-hydroxy-6-methyloxan-2-yl]oxy-10-[(2s,5s,6r)-5-(dimethylamino)-6-methyloxan-2-yl]oxy-4-hydroxy-5-methoxy-9,16-dimethyl-2-o Chemical compound O([C@H]1/C=C/C=C/C[C@@H](C)OC(=O)C[C@@H](O)[C@@H]([C@H]([C@@H](CC=O)C[C@H]1C)O[C@H]1[C@@H]([C@H]([C@H](O[C@@H]2O[C@@H](C)[C@H](O)[C@](C)(O)C2)[C@@H](C)O1)N(C)C)O)OC)[C@@H]1CC[C@H](N(C)C)[C@@H](C)O1 ACTOXUHEUCPTEW-BWHGAVFKSA-N 0.000 description 1
- XQCZBXHVTFVIFE-UHFFFAOYSA-N 2-amino-4-hydroxypyrimidine Chemical compound NC1=NC=CC(O)=N1 XQCZBXHVTFVIFE-UHFFFAOYSA-N 0.000 description 1
- VKNASXZDGZNEDA-UHFFFAOYSA-N 2-cyanoethyl 2-methylprop-2-enoate Chemical compound CC(=C)C(=O)OCCC#N VKNASXZDGZNEDA-UHFFFAOYSA-N 0.000 description 1
- SFPNZPQIIAJXGL-UHFFFAOYSA-N 2-ethoxyethyl 2-methylprop-2-enoate Chemical compound CCOCCOC(=O)C(C)=C SFPNZPQIIAJXGL-UHFFFAOYSA-N 0.000 description 1
- MUZDXNQOSGWMJJ-UHFFFAOYSA-N 2-methylprop-2-enoic acid;prop-2-enoic acid Chemical compound OC(=O)C=C.CC(=C)C(O)=O MUZDXNQOSGWMJJ-UHFFFAOYSA-N 0.000 description 1
- FUBFWTUFPGFHOJ-UHFFFAOYSA-N 2-nitrofuran Chemical class [O-][N+](=O)C1=CC=CO1 FUBFWTUFPGFHOJ-UHFFFAOYSA-N 0.000 description 1
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 1
- OLGCJTRSPFQVOV-UHFFFAOYSA-N 4-amino-n-(5,6-dimethoxypyrimidin-4-yl)benzenesulfonamide;5-[(3,4,5-trimethoxyphenyl)methyl]pyrimidine-2,4-diamine Chemical compound COC1=C(OC)C(OC)=CC(CC=2C(=NC(N)=NC=2)N)=C1.COC1=NC=NC(NS(=O)(=O)C=2C=CC(N)=CC=2)=C1OC OLGCJTRSPFQVOV-UHFFFAOYSA-N 0.000 description 1
- HIQIXEFWDLTDED-UHFFFAOYSA-N 4-hydroxy-1-piperidin-4-ylpyrrolidin-2-one Chemical compound O=C1CC(O)CN1C1CCNCC1 HIQIXEFWDLTDED-UHFFFAOYSA-N 0.000 description 1
- VDABVNMGKGUPEY-UHFFFAOYSA-N 6-carboxyfluorescein succinimidyl ester Chemical compound C=1C(O)=CC=C2C=1OC1=CC(O)=CC=C1C2(C1=C2)OC(=O)C1=CC=C2C(=O)ON1C(=O)CCC1=O VDABVNMGKGUPEY-UHFFFAOYSA-N 0.000 description 1
- WUWFMDMBOJLQIV-UHFFFAOYSA-N 7-(3-aminopyrrolidin-1-yl)-1-(2,4-difluorophenyl)-6-fluoro-4-oxo-1,4-dihydro-1,8-naphthyridine-3-carboxylic acid Chemical compound C1C(N)CCN1C(C(=C1)F)=NC2=C1C(=O)C(C(O)=O)=CN2C1=CC=C(F)C=C1F WUWFMDMBOJLQIV-UHFFFAOYSA-N 0.000 description 1
- MPORYQCGWFQFLA-ONPDANIMSA-N 7-[(7s)-7-amino-5-azaspiro[2.4]heptan-5-yl]-8-chloro-6-fluoro-1-[(1r,2s)-2-fluorocyclopropyl]-4-oxoquinoline-3-carboxylic acid;trihydrate Chemical compound O.O.O.C([C@H]1N)N(C=2C(=C3C(C(C(C(O)=O)=CN3[C@H]3[C@H](C3)F)=O)=CC=2F)Cl)CC11CC1.C([C@H]1N)N(C=2C(=C3C(C(C(C(O)=O)=CN3[C@H]3[C@H](C3)F)=O)=CC=2F)Cl)CC11CC1 MPORYQCGWFQFLA-ONPDANIMSA-N 0.000 description 1
- GSDSWSVVBLHKDQ-UHFFFAOYSA-N 9-fluoro-3-methyl-10-(4-methylpiperazin-1-yl)-7-oxo-2,3-dihydro-7H-[1,4]oxazino[2,3,4-ij]quinoline-6-carboxylic acid Chemical compound FC1=CC(C(C(C(O)=O)=C2)=O)=C3N2C(C)COC3=C1N1CCN(C)CC1 GSDSWSVVBLHKDQ-UHFFFAOYSA-N 0.000 description 1
- 108010024100 APOBEC Deaminases Proteins 0.000 description 1
- 102000015619 APOBEC Deaminases Human genes 0.000 description 1
- 108010004483 APOBEC-3G Deaminase Proteins 0.000 description 1
- 102000002797 APOBEC-3G Deaminase Human genes 0.000 description 1
- 244000215068 Acacia senegal Species 0.000 description 1
- 241000604451 Acidaminococcus Species 0.000 description 1
- 101000860090 Acidaminococcus sp. (strain BV3L6) CRISPR-associated endonuclease Cas12a Proteins 0.000 description 1
- 108060003345 Adrenergic Receptor Proteins 0.000 description 1
- 102000017910 Adrenergic receptor Human genes 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 229920000936 Agarose Polymers 0.000 description 1
- 241001135230 Alistipes putredinis Species 0.000 description 1
- 102000002226 Alkyl and Aryl Transferases Human genes 0.000 description 1
- 108010014722 Alkyl and Aryl Transferases Proteins 0.000 description 1
- 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 1
- 240000002339 Anredera cordifolia Species 0.000 description 1
- WZPBZJONDBGPKJ-UHFFFAOYSA-N Antibiotic SQ 26917 Natural products O=C1N(S(O)(=O)=O)C(C)C1NC(=O)C(=NOC(C)(C)C(O)=O)C1=CSC(N)=N1 WZPBZJONDBGPKJ-UHFFFAOYSA-N 0.000 description 1
- 235000003911 Arachis Nutrition 0.000 description 1
- 244000105624 Arachis hypogaea Species 0.000 description 1
- 241000203069 Archaea Species 0.000 description 1
- 241000193749 Bacillus coagulans Species 0.000 description 1
- 108020004513 Bacterial RNA Proteins 0.000 description 1
- 208000034309 Bacterial disease carrier Diseases 0.000 description 1
- 241001674039 Bacteroides acidifaciens Species 0.000 description 1
- 241000168635 Bacteroides barnesiae Species 0.000 description 1
- 241000217846 Bacteroides caccae Species 0.000 description 1
- 241000859775 Bacteroides caecicola Species 0.000 description 1
- 241000685477 Bacteroides caecigallinarum Species 0.000 description 1
- 241001032450 Bacteroides cellulosilyticus Species 0.000 description 1
- 241000801600 Bacteroides clarus Species 0.000 description 1
- 241001220439 Bacteroides coprocola Species 0.000 description 1
- 241000545821 Bacteroides coprophilus Species 0.000 description 1
- 241001631983 Bacteroides coprosuis Species 0.000 description 1
- 241001135322 Bacteroides eggerthii Species 0.000 description 1
- 241000337516 Bacteroides faecichinchillae Species 0.000 description 1
- 241000402140 Bacteroides finegoldii Species 0.000 description 1
- 241000801629 Bacteroides fluxus Species 0.000 description 1
- 241000514947 Bacteroides galacturonicus Species 0.000 description 1
- 241000168642 Bacteroides gallinarum Species 0.000 description 1
- 241001567982 Bacteroides graminisolvens Species 0.000 description 1
- 241001109645 Bacteroides helcogenes Species 0.000 description 1
- 241000047484 Bacteroides intestinalis Species 0.000 description 1
- 241000947128 Bacteroides luti Species 0.000 description 1
- 241001195773 Bacteroides massiliensis Species 0.000 description 1
- 241001122266 Bacteroides nordii Species 0.000 description 1
- 241000801630 Bacteroides oleiciplenus Species 0.000 description 1
- 241001135228 Bacteroides ovatus Species 0.000 description 1
- 241000828416 Bacteroides paurosaccharolyticus Species 0.000 description 1
- 241001220441 Bacteroides plebeius Species 0.000 description 1
- 241000859824 Bacteroides propionicifaciens Species 0.000 description 1
- 241000660102 Bacteroides reticulotermitis Species 0.000 description 1
- 241001652281 Bacteroides rodentium Species 0.000 description 1
- 241000168636 Bacteroides salanitronis Species 0.000 description 1
- 241001122267 Bacteroides salyersiae Species 0.000 description 1
- 241000911892 Bacteroides sartorii Species 0.000 description 1
- 241000204294 Bacteroides stercoris Species 0.000 description 1
- 241000606219 Bacteroides uniformis Species 0.000 description 1
- 241000115153 Bacteroides xylanisolvens Species 0.000 description 1
- 241000514942 Bacteroides xylanolyticus Species 0.000 description 1
- 241000605059 Bacteroidetes Species 0.000 description 1
- MGQLHRYJBWGORO-LLVKDONJSA-N Balofloxacin Chemical compound C1[C@H](NC)CCCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN(C3CC3)C2=C1OC MGQLHRYJBWGORO-LLVKDONJSA-N 0.000 description 1
- 239000005711 Benzoic acid Substances 0.000 description 1
- 241000219310 Beta vulgaris subsp. vulgaris Species 0.000 description 1
- 241000589969 Borreliella burgdorferi Species 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 102100027310 Bromodomain adjacent to zinc finger domain protein 1A Human genes 0.000 description 1
- 238000009631 Broth culture Methods 0.000 description 1
- 102100040399 C->U-editing enzyme APOBEC-2 Human genes 0.000 description 1
- 102100025074 C-C chemokine receptor-like 2 Human genes 0.000 description 1
- 102000017420 CD3 protein, epsilon/gamma/delta subunit Human genes 0.000 description 1
- 108050005493 CD3 protein, epsilon/gamma/delta subunit Proteins 0.000 description 1
- 108010040467 CRISPR-Associated Proteins Proteins 0.000 description 1
- 238000010356 CRISPR-Cas9 genome editing Methods 0.000 description 1
- 238000010453 CRISPR/Cas method Methods 0.000 description 1
- 101150018129 CSF2 gene Proteins 0.000 description 1
- 101150069031 CSN2 gene Proteins 0.000 description 1
- 229940045513 CTLA4 antagonist Drugs 0.000 description 1
- 102000000905 Cadherin Human genes 0.000 description 1
- 108050007957 Cadherin Proteins 0.000 description 1
- 241000589875 Campylobacter jejuni Species 0.000 description 1
- 241000282472 Canis lupus familiaris Species 0.000 description 1
- 108010065839 Capreomycin Proteins 0.000 description 1
- UKXFZNMZXWBSGH-UHFFFAOYSA-N Carbomycin A Natural products COC1C(OC2OC(C)C(OC3CC(C)(O)C(OC(=O)CC(C)C)C(C)O3)C(C2O)N(C)C)C(CC=O)CC(C)C(=O)C=CC4OC4CC(C)OC(=O)CC1C(=O)C UKXFZNMZXWBSGH-UHFFFAOYSA-N 0.000 description 1
- 239000004215 Carbon black (E152) Substances 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- 206010007556 Cardiac failure acute Diseases 0.000 description 1
- 208000006029 Cardiomegaly Diseases 0.000 description 1
- 241000700198 Cavia Species 0.000 description 1
- UQLLWWBDSUHNEB-CZUORRHYSA-N Cefaprin Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC(=O)C)C(O)=O)C(=O)CSC1=CC=NC=C1 UQLLWWBDSUHNEB-CZUORRHYSA-N 0.000 description 1
- QYQDKDWGWDOFFU-IUODEOHRSA-N Cefotiam Chemical compound CN(C)CCN1N=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CC=3N=C(N)SC=3)[C@H]2SC1 QYQDKDWGWDOFFU-IUODEOHRSA-N 0.000 description 1
- 108010051109 Cell-Penetrating Peptides Proteins 0.000 description 1
- 102000020313 Cell-Penetrating Peptides Human genes 0.000 description 1
- DQEFEBPAPFSJLV-UHFFFAOYSA-N Cellulose propionate Chemical compound CCC(=O)OCC1OC(OC(=O)CC)C(OC(=O)CC)C(OC(=O)CC)C1OC1C(OC(=O)CC)C(OC(=O)CC)C(OC(=O)CC)C(COC(=O)CC)O1 DQEFEBPAPFSJLV-UHFFFAOYSA-N 0.000 description 1
- 229930186147 Cephalosporin Natural products 0.000 description 1
- 108091092236 Chimeric RNA Proteins 0.000 description 1
- 241000700114 Chinchillidae Species 0.000 description 1
- 229920002101 Chitin Polymers 0.000 description 1
- 241000606161 Chlamydia Species 0.000 description 1
- 108020004705 Codon Proteins 0.000 description 1
- 208000003322 Coinfection Diseases 0.000 description 1
- 108010078777 Colistin Proteins 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010010264 Condition aggravated Diseases 0.000 description 1
- 241000699800 Cricetinae Species 0.000 description 1
- 229920002785 Croscarmellose sodium Polymers 0.000 description 1
- 101150074775 Csf1 gene Proteins 0.000 description 1
- 229920000858 Cyclodextrin Polymers 0.000 description 1
- 102220605874 Cytosolic arginine sensor for mTORC1 subunit 2_D10A_mutation Human genes 0.000 description 1
- DYDCUQKUCUHJBH-UWTATZPHSA-N D-Cycloserine Chemical compound N[C@@H]1CONC1=O DYDCUQKUCUHJBH-UWTATZPHSA-N 0.000 description 1
- DYDCUQKUCUHJBH-UHFFFAOYSA-N D-Cycloserine Natural products NC1CONC1=O DYDCUQKUCUHJBH-UHFFFAOYSA-N 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- RGHNJXZEOKUKBD-UHFFFAOYSA-N D-gluconic acid Natural products OCC(O)C(O)C(O)C(O)C(O)=O RGHNJXZEOKUKBD-UHFFFAOYSA-N 0.000 description 1
- 102100040262 DNA dC->dU-editing enzyme APOBEC-3B Human genes 0.000 description 1
- 102100040261 DNA dC->dU-editing enzyme APOBEC-3C Human genes 0.000 description 1
- 102100040264 DNA dC->dU-editing enzyme APOBEC-3D Human genes 0.000 description 1
- 102100040266 DNA dC->dU-editing enzyme APOBEC-3F Human genes 0.000 description 1
- 102100038050 DNA dC->dU-editing enzyme APOBEC-3H Human genes 0.000 description 1
- 101710082737 DNA dC->dU-editing enzyme APOBEC-3H Proteins 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 230000008836 DNA modification Effects 0.000 description 1
- 230000003682 DNA packaging effect Effects 0.000 description 1
- 230000004543 DNA replication Effects 0.000 description 1
- 230000007018 DNA scission Effects 0.000 description 1
- 102000052510 DNA-Binding Proteins Human genes 0.000 description 1
- 101710096438 DNA-binding protein Proteins 0.000 description 1
- 102210047350 DQA1*03:03 Human genes 0.000 description 1
- 102210047410 DQA1*05:05 Human genes 0.000 description 1
- 102210047213 DQB1*03:05 Human genes 0.000 description 1
- MQJKPEGWNLWLTK-UHFFFAOYSA-N Dapsone Chemical compound C1=CC(N)=CC=C1S(=O)(=O)C1=CC=C(N)C=C1 MQJKPEGWNLWLTK-UHFFFAOYSA-N 0.000 description 1
- 108010013198 Daptomycin Proteins 0.000 description 1
- FMTDIUIBLCQGJB-UHFFFAOYSA-N Demethylchlortetracyclin Natural products C1C2C(O)C3=C(Cl)C=CC(O)=C3C(=O)C2=C(O)C2(O)C1C(N(C)C)C(O)=C(C(N)=O)C2=O FMTDIUIBLCQGJB-UHFFFAOYSA-N 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 229920001353 Dextrin Polymers 0.000 description 1
- FEWJPZIEWOKRBE-JCYAYHJZSA-N Dextrotartaric acid Chemical compound OC(=O)[C@H](O)[C@@H](O)C(O)=O FEWJPZIEWOKRBE-JCYAYHJZSA-N 0.000 description 1
- 239000004338 Dichlorodifluoromethane Substances 0.000 description 1
- 206010061818 Disease progression Diseases 0.000 description 1
- JWCSIUVGFCSJCK-CAVRMKNVSA-N Disodium Moxalactam Chemical compound N([C@]1(OC)C(N2C(=C(CSC=3N(N=NN=3)C)CO[C@@H]21)C(O)=O)=O)C(=O)C(C(O)=O)C1=CC=C(O)C=C1 JWCSIUVGFCSJCK-CAVRMKNVSA-N 0.000 description 1
- 102100038191 Double-stranded RNA-specific editase 1 Human genes 0.000 description 1
- 101100490452 Drosophila melanogaster Adat1 gene Proteins 0.000 description 1
- LVGKNOAMLMIIKO-UHFFFAOYSA-N Elaidinsaeure-aethylester Natural products CCCCCCCCC=CCCCCCCCC(=O)OCC LVGKNOAMLMIIKO-UHFFFAOYSA-N 0.000 description 1
- 238000011510 Elispot assay Methods 0.000 description 1
- 108010042407 Endonucleases Proteins 0.000 description 1
- 241000194033 Enterococcus Species 0.000 description 1
- 241000709661 Enterovirus Species 0.000 description 1
- 239000004593 Epoxy Substances 0.000 description 1
- 241000283086 Equidae Species 0.000 description 1
- 241000283074 Equus asinus Species 0.000 description 1
- 241000588722 Escherichia Species 0.000 description 1
- 241001198387 Escherichia coli BL21(DE3) Species 0.000 description 1
- 241001646716 Escherichia coli K-12 Species 0.000 description 1
- IAYPIBMASNFSPL-UHFFFAOYSA-N Ethylene oxide Chemical compound C1CO1 IAYPIBMASNFSPL-UHFFFAOYSA-N 0.000 description 1
- 241000206602 Eukaryota Species 0.000 description 1
- 108700024394 Exon Proteins 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 206010016654 Fibrosis Diseases 0.000 description 1
- 238000000729 Fisher's exact test Methods 0.000 description 1
- 206010017533 Fungal infection Diseases 0.000 description 1
- IECPWNUMDGFDKC-UHFFFAOYSA-N Fusicsaeure Natural products C12C(O)CC3C(=C(CCC=C(C)C)C(O)=O)C(OC(C)=O)CC3(C)C1(C)CCC1C2(C)CCC(O)C1C IECPWNUMDGFDKC-UHFFFAOYSA-N 0.000 description 1
- JRZJKWGQFNTSRN-UHFFFAOYSA-N Geldanamycin Natural products C1C(C)CC(OC)C(O)C(C)C=C(C)C(OC(N)=O)C(OC)CCC=C(C)C(=O)NC2=CC(=O)C(OC)=C1C2=O JRZJKWGQFNTSRN-UHFFFAOYSA-N 0.000 description 1
- 108700039691 Genetic Promoter Regions Proteins 0.000 description 1
- 241000699694 Gerbillinae Species 0.000 description 1
- 229920002581 Glucomannan Polymers 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 108010015899 Glycopeptides Proteins 0.000 description 1
- 102000002068 Glycopeptides Human genes 0.000 description 1
- 229940113491 Glycosylase inhibitor Drugs 0.000 description 1
- 238000003794 Gram staining Methods 0.000 description 1
- AIJTTZAVMXIJGM-UHFFFAOYSA-N Grepafloxacin Chemical compound C1CNC(C)CN1C(C(=C1C)F)=CC2=C1C(=O)C(C(O)=O)=CN2C1CC1 AIJTTZAVMXIJGM-UHFFFAOYSA-N 0.000 description 1
- 108091029499 Group II intron Proteins 0.000 description 1
- 229920002907 Guar gum Polymers 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 108010045618 HLA-DQ7 antigen Proteins 0.000 description 1
- 108010078950 HLA-DQ9 antigen Proteins 0.000 description 1
- 102210049245 HLA-DQA1*05:01 Human genes 0.000 description 1
- 102210053890 HLA-DQB1*03:01 Human genes 0.000 description 1
- 229920002488 Hemicellulose Polymers 0.000 description 1
- 102100026122 High affinity immunoglobulin gamma Fc receptor I Human genes 0.000 description 1
- 101000937778 Homo sapiens Bromodomain adjacent to zinc finger domain protein 1A Proteins 0.000 description 1
- 101000964322 Homo sapiens C->U-editing enzyme APOBEC-2 Proteins 0.000 description 1
- 101000716068 Homo sapiens C-C chemokine receptor type 6 Proteins 0.000 description 1
- 101000964385 Homo sapiens DNA dC->dU-editing enzyme APOBEC-3B Proteins 0.000 description 1
- 101000964383 Homo sapiens DNA dC->dU-editing enzyme APOBEC-3C Proteins 0.000 description 1
- 101000964382 Homo sapiens DNA dC->dU-editing enzyme APOBEC-3D Proteins 0.000 description 1
- 101000964377 Homo sapiens DNA dC->dU-editing enzyme APOBEC-3F Proteins 0.000 description 1
- 101000742223 Homo sapiens Double-stranded RNA-specific editase 1 Proteins 0.000 description 1
- 101000913074 Homo sapiens High affinity immunoglobulin gamma Fc receptor I Proteins 0.000 description 1
- 101001046686 Homo sapiens Integrin alpha-M Proteins 0.000 description 1
- 101000830956 Homo sapiens Three-prime repair exonuclease 1 Proteins 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 239000004354 Hydroxyethyl cellulose Substances 0.000 description 1
- 229920000663 Hydroxyethyl cellulose Polymers 0.000 description 1
- UGQMRVRMYYASKQ-UHFFFAOYSA-N Hypoxanthine nucleoside Natural products OC1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 UGQMRVRMYYASKQ-UHFFFAOYSA-N 0.000 description 1
- 206010052768 Infectious myocarditis Diseases 0.000 description 1
- 102100022338 Integrin alpha-M Human genes 0.000 description 1
- 108091092195 Intron Proteins 0.000 description 1
- JUZNIMUFDBIJCM-ANEDZVCMSA-N Invanz Chemical compound O=C([C@H]1NC[C@H](C1)SC=1[C@H](C)[C@@H]2[C@H](C(N2C=1C(O)=O)=O)[C@H](O)C)NC1=CC=CC(C(O)=O)=C1 JUZNIMUFDBIJCM-ANEDZVCMSA-N 0.000 description 1
- MIFYHUACUWQUKT-UHFFFAOYSA-N Isopenicillin N Natural products OC(=O)C1C(C)(C)SC2C(NC(=O)CCCC(N)C(O)=O)C(=O)N21 MIFYHUACUWQUKT-UHFFFAOYSA-N 0.000 description 1
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 1
- XAGMUUZPGZWTRP-ZETCQYMHSA-N LSM-5745 Chemical compound C([C@@H](N1C2=C(C(C(C(O)=O)=C1)=O)C=C1F)C)OC2=C1C1(N)CC1 XAGMUUZPGZWTRP-ZETCQYMHSA-N 0.000 description 1
- 241001112693 Lachnospiraceae Species 0.000 description 1
- 239000004166 Lanolin Substances 0.000 description 1
- 239000005639 Lauric acid Substances 0.000 description 1
- 206010049694 Left Ventricular Dysfunction Diseases 0.000 description 1
- 241000029590 Leptotrichia wadei Species 0.000 description 1
- 101000860104 Leptotrichia wadei (strain F0279) CRISPR-associated endoribonuclease Cas13a Proteins 0.000 description 1
- 240000007472 Leucaena leucocephala Species 0.000 description 1
- 235000010643 Leucaena leucocephala Nutrition 0.000 description 1
- GSDSWSVVBLHKDQ-JTQLQIEISA-N Levofloxacin Chemical compound C([C@@H](N1C2=C(C(C(C(O)=O)=C1)=O)C=C1F)C)OC2=C1N1CCN(C)CC1 GSDSWSVVBLHKDQ-JTQLQIEISA-N 0.000 description 1
- 108010028921 Lipopeptides Proteins 0.000 description 1
- 229920002774 Maltodextrin Polymers 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 201000009906 Meningitis Diseases 0.000 description 1
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 description 1
- AFVFQIVMOAPDHO-UHFFFAOYSA-N Methanesulfonic acid Chemical compound CS(O)(=O)=O AFVFQIVMOAPDHO-UHFFFAOYSA-N 0.000 description 1
- RJQXTJLFIWVMTO-TYNCELHUSA-N Methicillin Chemical compound COC1=CC=CC(OC)=C1C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 RJQXTJLFIWVMTO-TYNCELHUSA-N 0.000 description 1
- DMUAPQTXSSNEDD-QALJCMCCSA-N Midecamycin Chemical compound C1[C@](O)(C)[C@@H](OC(=O)CC)[C@H](C)O[C@H]1O[C@H]1[C@H](N(C)C)[C@@H](O)[C@H](O[C@@H]2[C@H]([C@H](OC(=O)CC)CC(=O)O[C@H](C)C/C=C/C=C/[C@H](O)[C@H](C)C[C@@H]2CC=O)OC)O[C@@H]1C DMUAPQTXSSNEDD-QALJCMCCSA-N 0.000 description 1
- 101100219625 Mus musculus Casd1 gene Proteins 0.000 description 1
- 241000282339 Mustela Species 0.000 description 1
- 208000031888 Mycoses Diseases 0.000 description 1
- 102000005604 Myosin Heavy Chains Human genes 0.000 description 1
- 108010084498 Myosin Heavy Chains Proteins 0.000 description 1
- FXHOOIRPVKKKFG-UHFFFAOYSA-N N,N-Dimethylacetamide Chemical compound CN(C)C(C)=O FXHOOIRPVKKKFG-UHFFFAOYSA-N 0.000 description 1
- GWBPFRGXNGPPMF-UHFFFAOYSA-N N-[4-[(4-nitrophenyl)sulfamoyl]phenyl]acetamide Chemical compound C1=CC(NC(=O)C)=CC=C1S(=O)(=O)NC1=CC=C([N+]([O-])=O)C=C1 GWBPFRGXNGPPMF-UHFFFAOYSA-N 0.000 description 1
- 241000588653 Neisseria Species 0.000 description 1
- 229930193140 Neomycin Natural products 0.000 description 1
- QSDSSSQWVNLFIG-UHFFFAOYSA-N Neosporin Natural products CC(O)CC1=C(OC)C(=O)C2=CC(O)=C3OCOC4=C(O)C=C5C6=C4C3=C2C1=C6C(CC(C)O)=C(OC)C5=O QSDSSSQWVNLFIG-UHFFFAOYSA-N 0.000 description 1
- 101100385413 Neurospora crassa (strain ATCC 24698 / 74-OR23-1A / CBS 708.71 / DSM 1257 / FGSC 987) csm-3 gene Proteins 0.000 description 1
- 239000004104 Oleandomycin Substances 0.000 description 1
- RZPAKFUAFGMUPI-UHFFFAOYSA-N Oleandomycin Natural products O1C(C)C(O)C(OC)CC1OC1C(C)C(=O)OC(C)C(C)C(O)C(C)C(=O)C2(OC2)CC(C)C(OC2C(C(CC(C)O2)N(C)C)O)C1C RZPAKFUAFGMUPI-UHFFFAOYSA-N 0.000 description 1
- KYGZCKSPAKDVKC-UHFFFAOYSA-N Oxolinic acid Chemical compound C1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC2=C1OCO2 KYGZCKSPAKDVKC-UHFFFAOYSA-N 0.000 description 1
- 239000004100 Oxytetracycline Substances 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 241000030714 Parabacteroides goldsteinii Species 0.000 description 1
- 229930040373 Paraformaldehyde Natural products 0.000 description 1
- UOZODPSAJZTQNH-UHFFFAOYSA-N Paromomycin II Natural products NC1C(O)C(O)C(CN)OC1OC1C(O)C(OC2C(C(N)CC(N)C2O)OC2C(C(O)C(O)C(CO)O2)N)OC1CO UOZODPSAJZTQNH-UHFFFAOYSA-N 0.000 description 1
- 208000031481 Pathologic Constriction Diseases 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- XVASOOUVMJAZNJ-MBNYWOFBSA-N Penicillin K Chemical compound S1C(C)(C)[C@H](C(O)=O)N2C(=O)[C@@H](NC(=O)CCCCCCC)[C@H]21 XVASOOUVMJAZNJ-MBNYWOFBSA-N 0.000 description 1
- 229930195708 Penicillin V Natural products 0.000 description 1
- 108091093037 Peptide nucleic acid Proteins 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 208000037581 Persistent Infection Diseases 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 229920000954 Polyglycolide Polymers 0.000 description 1
- 108010093965 Polymyxin B Proteins 0.000 description 1
- 108010040201 Polymyxins Proteins 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 239000004793 Polystyrene Substances 0.000 description 1
- 239000004372 Polyvinyl alcohol Substances 0.000 description 1
- 241001135223 Prevotella melaninogenica Species 0.000 description 1
- 241001135262 Prevotella oris Species 0.000 description 1
- 102000001253 Protein Kinase Human genes 0.000 description 1
- 102000055027 Protein Methyltransferases Human genes 0.000 description 1
- 108700040121 Protein Methyltransferases Proteins 0.000 description 1
- 241000192142 Proteobacteria Species 0.000 description 1
- 208000010362 Protozoan Infections Diseases 0.000 description 1
- PWNMXPDKBYZCOO-UHFFFAOYSA-N Prulifloxacin Chemical compound C1=C2N3C(C)SC3=C(C(O)=O)C(=O)C2=CC(F)=C1N(CC1)CCN1CC=1OC(=O)OC=1C PWNMXPDKBYZCOO-UHFFFAOYSA-N 0.000 description 1
- 241001495182 Pseudobacteroides cellulosolvens Species 0.000 description 1
- 101100047461 Rattus norvegicus Trpm8 gene Proteins 0.000 description 1
- 108020004511 Recombinant DNA Proteins 0.000 description 1
- 229920000294 Resistant starch Polymers 0.000 description 1
- URWAJWIAIPFPJE-UHFFFAOYSA-N Rickamicin Natural products O1CC(O)(C)C(NC)C(O)C1OC1C(O)C(OC2C(CC=C(CN)O2)N)C(N)CC1N URWAJWIAIPFPJE-UHFFFAOYSA-N 0.000 description 1
- NJCJBUHJQLFDSW-UHFFFAOYSA-N Rufloxacin Chemical compound C1CN(C)CCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN3CCSC1=C32 NJCJBUHJQLFDSW-UHFFFAOYSA-N 0.000 description 1
- 241000134861 Ruminococcus sp. Species 0.000 description 1
- 241000235342 Saccharomycetes Species 0.000 description 1
- 108091058545 Secretory proteins Proteins 0.000 description 1
- 102000040739 Secretory proteins Human genes 0.000 description 1
- 108010071390 Serum Albumin Proteins 0.000 description 1
- 102000007562 Serum Albumin Human genes 0.000 description 1
- 229920001800 Shellac Polymers 0.000 description 1
- 108020004682 Single-Stranded DNA Proteins 0.000 description 1
- 229930192786 Sisomicin Natural products 0.000 description 1
- 229920002125 Sokalan® Polymers 0.000 description 1
- HVUMOYIDDBPOLL-XWVZOOPGSA-N Sorbitan monostearate Chemical compound CCCCCCCCCCCCCCCCCC(=O)OC[C@@H](O)[C@H]1OC[C@H](O)[C@H]1O HVUMOYIDDBPOLL-XWVZOOPGSA-N 0.000 description 1
- 239000004147 Sorbitan trioleate Substances 0.000 description 1
- PRXRUNOAOLTIEF-ADSICKODSA-N Sorbitan trioleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OC[C@@H](OC(=O)CCCCCCC\C=C/CCCCCCCC)[C@H]1OC[C@H](O)[C@H]1OC(=O)CCCCCCC\C=C/CCCCCCCC PRXRUNOAOLTIEF-ADSICKODSA-N 0.000 description 1
- 239000004187 Spiramycin Substances 0.000 description 1
- 241000194017 Streptococcus Species 0.000 description 1
- 241000194019 Streptococcus mutans Species 0.000 description 1
- 101000910035 Streptococcus pyogenes serotype M1 CRISPR-associated endonuclease Cas9/Csn1 Proteins 0.000 description 1
- 101000910045 Streptococcus thermophilus (strain ATCC BAA-491 / LMD-9) CRISPR-associated endonuclease Cas9 2 Proteins 0.000 description 1
- KDYFGRWQOYBRFD-UHFFFAOYSA-N Succinic acid Natural products OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 1
- 235000021536 Sugar beet Nutrition 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 230000006044 T cell activation Effects 0.000 description 1
- 108091008874 T cell receptors Proteins 0.000 description 1
- 102000016266 T-Cell Antigen Receptors Human genes 0.000 description 1
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 1
- 108010053950 Teicoplanin Proteins 0.000 description 1
- WKDDRNSBRWANNC-UHFFFAOYSA-N Thienamycin Natural products C1C(SCCN)=C(C(O)=O)N2C(=O)C(C(O)C)C21 WKDDRNSBRWANNC-UHFFFAOYSA-N 0.000 description 1
- 102100024855 Three-prime repair exonuclease 1 Human genes 0.000 description 1
- HJLSLZFTEKNLFI-UHFFFAOYSA-N Tinidazole Chemical compound CCS(=O)(=O)CCN1C(C)=NC=C1[N+]([O-])=O HJLSLZFTEKNLFI-UHFFFAOYSA-N 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 241000223109 Trypanosoma cruzi Species 0.000 description 1
- 239000004182 Tylosin Substances 0.000 description 1
- 229930194936 Tylosin Natural products 0.000 description 1
- 108091023045 Untranslated Region Proteins 0.000 description 1
- 102000006943 Uracil-DNA Glycosidase Human genes 0.000 description 1
- 108010072685 Uracil-DNA Glycosidase Proteins 0.000 description 1
- 101710172430 Uracil-DNA glycosylase inhibitor Proteins 0.000 description 1
- 206010052428 Wound Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 229920002000 Xyloglucan Polymers 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 229920002494 Zein Polymers 0.000 description 1
- FQVHOULQCKDUCY-OGHXVOSASA-N [(2s,3s,4r,6s)-6-[(2r,3s,4r,5r,6s)-6-[[(1s,3r,7r,8s,9s,10r,12r,14e,16s)-7-acetyloxy-8-methoxy-3,12-dimethyl-5,13-dioxo-10-(2-oxoethyl)-4,17-dioxabicyclo[14.1.0]heptadec-14-en-9-yl]oxy]-4-(dimethylamino)-5-hydroxy-2-methyloxan-3-yl]oxy-4-hydroxy-2,4-dimeth Chemical compound O([C@@H]1[C@@H](C)O[C@H]([C@@H]([C@H]1N(C)C)O)O[C@H]1[C@@H](CC=O)C[C@@H](C)C(=O)/C=C/[C@@H]2O[C@H]2C[C@@H](C)OC(=O)C[C@H]([C@@H]1OC)OC(C)=O)[C@H]1C[C@@](C)(O)[C@@H](OC(=O)CC(C)C)[C@H](C)O1 FQVHOULQCKDUCY-OGHXVOSASA-N 0.000 description 1
- ZWBTYMGEBZUQTK-PVLSIAFMSA-N [(7S,9E,11S,12R,13S,14R,15R,16R,17S,18S,19E,21Z)-2,15,17,32-tetrahydroxy-11-methoxy-3,7,12,14,16,18,22-heptamethyl-1'-(2-methylpropyl)-6,23-dioxospiro[8,33-dioxa-24,27,29-triazapentacyclo[23.6.1.14,7.05,31.026,30]tritriaconta-1(32),2,4,9,19,21,24,26,30-nonaene-28,4'-piperidine]-13-yl] acetate Chemical compound CO[C@H]1\C=C\O[C@@]2(C)Oc3c(C2=O)c2c4NC5(CCN(CC(C)C)CC5)N=c4c(=NC(=O)\C(C)=C/C=C/[C@H](C)[C@H](O)[C@@H](C)[C@@H](O)[C@@H](C)[C@H](OC(C)=O)[C@@H]1C)c(O)c2c(O)c3C ZWBTYMGEBZUQTK-PVLSIAFMSA-N 0.000 description 1
- UGXQOOQUZRUVSS-ZZXKWVIFSA-N [5-[3,5-dihydroxy-2-(1,3,4-trihydroxy-5-oxopentan-2-yl)oxyoxan-4-yl]oxy-3,4-dihydroxyoxolan-2-yl]methyl (e)-3-(4-hydroxyphenyl)prop-2-enoate Chemical compound OC1C(OC(CO)C(O)C(O)C=O)OCC(O)C1OC1C(O)C(O)C(COC(=O)\C=C\C=2C=CC(O)=CC=2)O1 UGXQOOQUZRUVSS-ZZXKWVIFSA-N 0.000 description 1
- 241001531189 [Eubacterium] siraeum Species 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- PENDGIOBPJLVBT-HMMOOPTJSA-N abt-773 Chemical compound O([C@@H]1[C@@H](C)C(=O)[C@@H](C)C(=O)O[C@@H]([C@]2(OC(=O)N[C@@H]2[C@@H](C)C(=O)[C@H](C)C[C@]1(C)OC\C=C\C=1C=C2C=CC=CC2=NC=1)C)CC)[C@@H]1O[C@H](C)C[C@H](N(C)C)[C@H]1O PENDGIOBPJLVBT-HMMOOPTJSA-N 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 235000011054 acetic acid Nutrition 0.000 description 1
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 229950008644 adicillin Drugs 0.000 description 1
- 239000001361 adipic acid Substances 0.000 description 1
- 235000011037 adipic acid Nutrition 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 229960000919 alatrofloxacin Drugs 0.000 description 1
- 230000007815 allergy Effects 0.000 description 1
- 229950008560 almecillin Drugs 0.000 description 1
- 238000005267 amalgamation Methods 0.000 description 1
- 230000001668 ameliorated effect Effects 0.000 description 1
- 229960004821 amikacin Drugs 0.000 description 1
- LKCWBDHBTVXHDL-RMDFUYIESA-N amikacin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O1)O)NC(=O)[C@@H](O)CCN)[C@H]1O[C@H](CN)[C@@H](O)[C@H](O)[C@H]1O LKCWBDHBTVXHDL-RMDFUYIESA-N 0.000 description 1
- 229960003022 amoxicillin Drugs 0.000 description 1
- LSQZJLSUYDQPKJ-NJBDSQKTSA-N amoxicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=C(O)C=C1 LSQZJLSUYDQPKJ-NJBDSQKTSA-N 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 150000008064 anhydrides Chemical class 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 230000000078 anti-malarial effect Effects 0.000 description 1
- 230000000845 anti-microbial effect Effects 0.000 description 1
- 230000002141 anti-parasite Effects 0.000 description 1
- 230000000561 anti-psychotic effect Effects 0.000 description 1
- 230000000798 anti-retroviral effect Effects 0.000 description 1
- 230000000692 anti-sense effect Effects 0.000 description 1
- 238000011394 anticancer treatment Methods 0.000 description 1
- 230000030741 antigen processing and presentation Effects 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 229920000617 arabinoxylan Polymers 0.000 description 1
- VLAXZGHHBIJLAD-UHFFFAOYSA-N arsphenamine Chemical compound [Cl-].[Cl-].C1=C(O)C([NH3+])=CC([As]=[As]C=2C=C([NH3+])C(O)=CC=2)=C1 VLAXZGHHBIJLAD-UHFFFAOYSA-N 0.000 description 1
- 229940003446 arsphenamine Drugs 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 208000037979 autoimmune inflammatory disease Diseases 0.000 description 1
- 230000006472 autoimmune response Effects 0.000 description 1
- 230000003816 axenic effect Effects 0.000 description 1
- 229960004099 azithromycin Drugs 0.000 description 1
- MQTOSJVFKKJCRP-BICOPXKESA-N azithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)N(C)C[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 MQTOSJVFKKJCRP-BICOPXKESA-N 0.000 description 1
- WZPBZJONDBGPKJ-VEHQQRBSSA-N aztreonam Chemical compound O=C1N(S([O-])(=O)=O)[C@@H](C)[C@@H]1NC(=O)C(=N/OC(C)(C)C(O)=O)\C1=CSC([NH3+])=N1 WZPBZJONDBGPKJ-VEHQQRBSSA-N 0.000 description 1
- 229960003644 aztreonam Drugs 0.000 description 1
- 229960002699 bacampicillin Drugs 0.000 description 1
- PFOLLRNADZZWEX-FFGRCDKISA-N bacampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)[C@H](C(S3)(C)C)C(=O)OC(C)OC(=O)OCC)=CC=CC=C1 PFOLLRNADZZWEX-FFGRCDKISA-N 0.000 description 1
- 229950000805 balofloxacin Drugs 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229960000686 benzalkonium chloride Drugs 0.000 description 1
- 235000010233 benzoic acid Nutrition 0.000 description 1
- CADWTSSKOVRVJC-UHFFFAOYSA-N benzyl(dimethyl)azanium;chloride Chemical compound [Cl-].C[NH+](C)CC1=CC=CC=C1 CADWTSSKOVRVJC-UHFFFAOYSA-N 0.000 description 1
- 239000003782 beta lactam antibiotic agent Substances 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 102000014992 beta1-adrenergic receptor activity proteins Human genes 0.000 description 1
- 108040006808 beta1-adrenergic receptor activity proteins Proteins 0.000 description 1
- 210000000941 bile Anatomy 0.000 description 1
- 239000003833 bile salt Substances 0.000 description 1
- 229940093761 bile salts Drugs 0.000 description 1
- 230000001851 biosynthetic effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- KQNZDYYTLMIZCT-KQPMLPITSA-N brefeldin A Chemical compound O[C@@H]1\C=C\C(=O)O[C@@H](C)CCC\C=C\[C@@H]2C[C@H](O)C[C@H]21 KQNZDYYTLMIZCT-KQPMLPITSA-N 0.000 description 1
- JUMGSHROWPPKFX-UHFFFAOYSA-N brefeldin-A Natural products CC1CCCC=CC2(C)CC(O)CC2(C)C(O)C=CC(=O)O1 JUMGSHROWPPKFX-UHFFFAOYSA-N 0.000 description 1
- KDYFGRWQOYBRFD-NUQCWPJISA-N butanedioic acid Chemical compound O[14C](=O)CC[14C](O)=O KDYFGRWQOYBRFD-NUQCWPJISA-N 0.000 description 1
- 238000004422 calculation algorithm Methods 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 239000004204 candelilla wax Substances 0.000 description 1
- 235000013868 candelilla wax Nutrition 0.000 description 1
- 229940073532 candelilla wax Drugs 0.000 description 1
- 229960004602 capreomycin Drugs 0.000 description 1
- 229940041011 carbapenems Drugs 0.000 description 1
- 229960003669 carbenicillin Drugs 0.000 description 1
- FPPNZSSZRUTDAP-UWFZAAFLSA-N carbenicillin Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)C(C(O)=O)C1=CC=CC=C1 FPPNZSSZRUTDAP-UWFZAAFLSA-N 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 239000011545 carbonate/bicarbonate buffer Substances 0.000 description 1
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 1
- 125000002057 carboxymethyl group Chemical group [H]OC(=O)C([H])([H])[*] 0.000 description 1
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 230000003683 cardiac damage Effects 0.000 description 1
- 210000004413 cardiac myocyte Anatomy 0.000 description 1
- 230000001426 cardiotropic effect Effects 0.000 description 1
- 235000010418 carrageenan Nutrition 0.000 description 1
- 229920001525 carrageenan Polymers 0.000 description 1
- 239000000679 carrageenan Substances 0.000 description 1
- 229940113118 carrageenan Drugs 0.000 description 1
- 101150055766 cat gene Proteins 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- 229920006317 cationic polymer Polymers 0.000 description 1
- 230000001364 causal effect Effects 0.000 description 1
- 229960003972 cefacetrile Drugs 0.000 description 1
- RRYMAQUWDLIUPV-BXKDBHETSA-N cefacetrile Chemical compound S1CC(COC(=O)C)=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CC#N)[C@@H]12 RRYMAQUWDLIUPV-BXKDBHETSA-N 0.000 description 1
- 229960005361 cefaclor Drugs 0.000 description 1
- QYIYFLOTGYLRGG-GPCCPHFNSA-N cefaclor Chemical compound C1([C@H](C(=O)N[C@@H]2C(N3C(=C(Cl)CS[C@@H]32)C(O)=O)=O)N)=CC=CC=C1 QYIYFLOTGYLRGG-GPCCPHFNSA-N 0.000 description 1
- 229960004841 cefadroxil Drugs 0.000 description 1
- NBFNMSULHIODTC-CYJZLJNKSA-N cefadroxil monohydrate Chemical compound O.C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)C)C(O)=O)=CC=C(O)C=C1 NBFNMSULHIODTC-CYJZLJNKSA-N 0.000 description 1
- FUBBGQLTSCSAON-PBFPGSCMSA-N cefaloglycin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)COC(=O)C)C(O)=O)=CC=CC=C1 FUBBGQLTSCSAON-PBFPGSCMSA-N 0.000 description 1
- 229950004030 cefaloglycin Drugs 0.000 description 1
- 229950005258 cefalonium Drugs 0.000 description 1
- GOFCPYKUMJBHBH-RHSMWYFYSA-N cefaloram Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC(=O)C)C(O)=O)C(=O)CC1=CC=CC=C1 GOFCPYKUMJBHBH-RHSMWYFYSA-N 0.000 description 1
- 229950001373 cefaloram Drugs 0.000 description 1
- 229960003866 cefaloridine Drugs 0.000 description 1
- CZTQZXZIADLWOZ-CRAIPNDOSA-N cefaloridine Chemical compound O=C([C@@H](NC(=O)CC=1SC=CC=1)[C@H]1SC2)N1C(C(=O)[O-])=C2C[N+]1=CC=CC=C1 CZTQZXZIADLWOZ-CRAIPNDOSA-N 0.000 description 1
- 229960000603 cefalotin Drugs 0.000 description 1
- 229950000042 cefaparole Drugs 0.000 description 1
- 229960004350 cefapirin Drugs 0.000 description 1
- 229960002420 cefatrizine Drugs 0.000 description 1
- UOCJDOLVGGIYIQ-PBFPGSCMSA-N cefatrizine Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)[C@H](N)C=2C=CC(O)=CC=2)CC=1CSC=1C=NNN=1 UOCJDOLVGGIYIQ-PBFPGSCMSA-N 0.000 description 1
- HGXLJRWXCXSEJO-GMSGAONNSA-N cefazaflur Chemical compound CN1N=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CSC(F)(F)F)[C@H]2SC1 HGXLJRWXCXSEJO-GMSGAONNSA-N 0.000 description 1
- 229950004359 cefazaflur Drugs 0.000 description 1
- 229960005312 cefazedone Drugs 0.000 description 1
- VTLCNEGVSVJLDN-MLGOLLRUSA-N cefazedone Chemical compound S1C(C)=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CN3C=C(Cl)C(=O)C(Cl)=C3)[C@H]2SC1 VTLCNEGVSVJLDN-MLGOLLRUSA-N 0.000 description 1
- 229960001139 cefazolin Drugs 0.000 description 1
- MLYYVTUWGNIJIB-BXKDBHETSA-N cefazolin Chemical compound S1C(C)=NN=C1SCC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)CN3N=NN=C3)[C@H]2SC1 MLYYVTUWGNIJIB-BXKDBHETSA-N 0.000 description 1
- 229960001817 cefbuperazone Drugs 0.000 description 1
- SMSRCGPDNDCXFR-CYWZMYCQSA-N cefbuperazone Chemical compound O=C1C(=O)N(CC)CCN1C(=O)N[C@H]([C@H](C)O)C(=O)N[C@]1(OC)C(=O)N2C(C(O)=O)=C(CSC=3N(N=NN=3)C)CS[C@@H]21 SMSRCGPDNDCXFR-CYWZMYCQSA-N 0.000 description 1
- 229950002706 cefcanel Drugs 0.000 description 1
- 229960002966 cefcapene Drugs 0.000 description 1
- HJJRIJDTIPFROI-NVKITGPLSA-N cefcapene Chemical compound N([C@@H]1C(N2C(=C(COC(N)=O)CS[C@@H]21)C(O)=O)=O)C(=O)\C(=C/CC)C1=CSC(N)=N1 HJJRIJDTIPFROI-NVKITGPLSA-N 0.000 description 1
- JUVHVMCKLDZLGN-TVNFHGJBSA-N cefclidin Chemical compound N([C@@H]1C(N2C(=C(C[N+]34CCC(CC3)(CC4)C(N)=O)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=NSC(N)=N1 JUVHVMCKLDZLGN-TVNFHGJBSA-N 0.000 description 1
- HOGISBSFFHDTRM-GHXIOONMSA-N cefdaloxime Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC)C(O)=O)C(=O)C(=N/O)\C1=CSC(N)=N1 HOGISBSFFHDTRM-GHXIOONMSA-N 0.000 description 1
- 229950006550 cefdaloxime Drugs 0.000 description 1
- 229960003719 cefdinir Drugs 0.000 description 1
- RTXOFQZKPXMALH-GHXIOONMSA-N cefdinir Chemical compound S1C(N)=NC(C(=N\O)\C(=O)N[C@@H]2C(N3C(=C(C=C)CS[C@@H]32)C(O)=O)=O)=C1 RTXOFQZKPXMALH-GHXIOONMSA-N 0.000 description 1
- 229960004069 cefditoren Drugs 0.000 description 1
- KMIPKYQIOVAHOP-YLGJWRNMSA-N cefditoren Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1\C=C/C=1SC=NC=1C KMIPKYQIOVAHOP-YLGJWRNMSA-N 0.000 description 1
- 229950007281 cefedrolor Drugs 0.000 description 1
- 229950009347 cefempidone Drugs 0.000 description 1
- HVFLCNVBZFFHBT-ZKDACBOMSA-N cefepime Chemical compound S([C@@H]1[C@@H](C(N1C=1C([O-])=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1C[N+]1(C)CCCC1 HVFLCNVBZFFHBT-ZKDACBOMSA-N 0.000 description 1
- 229960002100 cefepime Drugs 0.000 description 1
- 229960004041 cefetamet Drugs 0.000 description 1
- MQLRYUCJDNBWMV-GHXIOONMSA-N cefetamet Chemical compound N([C@@H]1C(N2C(=C(C)CS[C@@H]21)C(O)=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 MQLRYUCJDNBWMV-GHXIOONMSA-N 0.000 description 1
- 229950003098 cefetrizole Drugs 0.000 description 1
- 229950007546 cefivitril Drugs 0.000 description 1
- 229960002129 cefixime Drugs 0.000 description 1
- OKBVVJOGVLARMR-QSWIMTSFSA-N cefixime Chemical compound S1C(N)=NC(C(=N\OCC(O)=O)\C(=O)N[C@@H]2C(N3C(=C(C=C)CS[C@@H]32)C(O)=O)=O)=C1 OKBVVJOGVLARMR-QSWIMTSFSA-N 0.000 description 1
- XAKKNLNAJBNLPC-MAYKBZFQSA-N cefluprenam Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)/C=C/C[N+](C)(CC)CC(N)=O)C([O-])=O)C(=O)C(=N/OCF)\C1=NSC(N)=N1 XAKKNLNAJBNLPC-MAYKBZFQSA-N 0.000 description 1
- 229950001334 cefluprenam Drugs 0.000 description 1
- UEQVTKSAEXANEZ-YCRCPZNHSA-N cefmatilen Chemical compound S1C(N)=NC(C(=N\O)\C(=O)N[C@@H]2C(N3C(=C(SCSC4=NNN=C4)CS[C@@H]32)C(O)=O)=O)=C1 UEQVTKSAEXANEZ-YCRCPZNHSA-N 0.000 description 1
- 229950008727 cefmatilen Drugs 0.000 description 1
- 229960003791 cefmenoxime Drugs 0.000 description 1
- HJJDBAOLQAWBMH-YCRCPZNHSA-N cefmenoxime Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NN=NN1C HJJDBAOLQAWBMH-YCRCPZNHSA-N 0.000 description 1
- 229960003585 cefmetazole Drugs 0.000 description 1
- SNBUBQHDYVFSQF-HIFRSBDPSA-N cefmetazole Chemical compound S([C@@H]1[C@@](C(N1C=1C(O)=O)=O)(NC(=O)CSCC#N)OC)CC=1CSC1=NN=NN1C SNBUBQHDYVFSQF-HIFRSBDPSA-N 0.000 description 1
- 229960002025 cefminox Drugs 0.000 description 1
- JSDXOWVAHXDYCU-VXSYNFHWSA-N cefminox Chemical compound S([C@@H]1[C@@](C(N1C=1C(O)=O)=O)(NC(=O)CSC[C@@H](N)C(O)=O)OC)CC=1CSC1=NN=NN1C JSDXOWVAHXDYCU-VXSYNFHWSA-N 0.000 description 1
- 229960001958 cefodizime Drugs 0.000 description 1
- XDZKBRJLTGRPSS-BGZQYGJUSA-N cefodizime Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NC(C)=C(CC(O)=O)S1 XDZKBRJLTGRPSS-BGZQYGJUSA-N 0.000 description 1
- 229960004489 cefonicid Drugs 0.000 description 1
- DYAIAHUQIPBDIP-AXAPSJFSSA-N cefonicid Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)[C@H](O)C=2C=CC=CC=2)CC=1CSC1=NN=NN1CS(O)(=O)=O DYAIAHUQIPBDIP-AXAPSJFSSA-N 0.000 description 1
- 229960004682 cefoperazone Drugs 0.000 description 1
- GCFBRXLSHGKWDP-XCGNWRKASA-N cefoperazone Chemical compound O=C1C(=O)N(CC)CCN1C(=O)N[C@H](C=1C=CC(O)=CC=1)C(=O)N[C@@H]1C(=O)N2C(C(O)=O)=C(CSC=3N(N=NN=3)C)CS[C@@H]21 GCFBRXLSHGKWDP-XCGNWRKASA-N 0.000 description 1
- ZINFAXPQMLDEEJ-GFVOIPPFSA-N cefoselis Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CN1C=CC(=N)N1CCO ZINFAXPQMLDEEJ-GFVOIPPFSA-N 0.000 description 1
- 229950001580 cefoselis Drugs 0.000 description 1
- 229960004261 cefotaxime Drugs 0.000 description 1
- AZZMGZXNTDTSME-JUZDKLSSSA-M cefotaxime sodium Chemical compound [Na+].N([C@@H]1C(N2C(=C(COC(C)=O)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 AZZMGZXNTDTSME-JUZDKLSSSA-M 0.000 description 1
- 229960001242 cefotiam Drugs 0.000 description 1
- 229960003391 cefovecin Drugs 0.000 description 1
- ZJGQFXVQDVCVOK-MSUXKOGISA-N cefovecin Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)/C(=N/OC)C=2N=C(N)SC=2)CC=1[C@@H]1CCCO1 ZJGQFXVQDVCVOK-MSUXKOGISA-N 0.000 description 1
- 229950002823 cefoxazole Drugs 0.000 description 1
- QDUIJCOKQCCXQY-WHJQOFBOSA-N cefozopran Chemical compound N([C@@H]1C(N2C(=C(CN3C4=CC=CN=[N+]4C=C3)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=NSC(N)=N1 QDUIJCOKQCCXQY-WHJQOFBOSA-N 0.000 description 1
- 229960002642 cefozopran Drugs 0.000 description 1
- LNZMRLHZGOBKAN-KAWPREARSA-N cefpimizole Chemical compound N1=CNC(C(=O)N[C@@H](C(=O)N[C@@H]2C(N3C(=C(C[N+]=4C=CC(CCS(O)(=O)=O)=CC=4)CS[C@@H]32)C([O-])=O)=O)C=2C=CC=CC=2)=C1C(=O)O LNZMRLHZGOBKAN-KAWPREARSA-N 0.000 description 1
- 229950004036 cefpimizole Drugs 0.000 description 1
- DKOQGJHPHLTOJR-WHRDSVKCSA-N cefpirome Chemical compound N([C@@H]1C(N2C(=C(C[N+]=3C=4CCCC=4C=CC=3)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 DKOQGJHPHLTOJR-WHRDSVKCSA-N 0.000 description 1
- 229960000466 cefpirome Drugs 0.000 description 1
- 229960005090 cefpodoxime Drugs 0.000 description 1
- WYUSVOMTXWRGEK-HBWVYFAYSA-N cefpodoxime Chemical compound N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC)C(O)=O)C(=O)C(=N/OC)\C1=CSC(N)=N1 WYUSVOMTXWRGEK-HBWVYFAYSA-N 0.000 description 1
- 229960002580 cefprozil Drugs 0.000 description 1
- 229950009592 cefquinome Drugs 0.000 description 1
- 229960002588 cefradine Drugs 0.000 description 1
- 229950003685 cefrotil Drugs 0.000 description 1
- 229960003844 cefroxadine Drugs 0.000 description 1
- RDMOROXKXONCAL-UEKVPHQBSA-N cefroxadine Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)OC)C(O)=O)=CCC=CC1 RDMOROXKXONCAL-UEKVPHQBSA-N 0.000 description 1
- OFKRKCHCYWQZLY-XHBSWPGZSA-N cefsumide Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)C)C(O)=O)=CC=CC(NS(C)(=O)=O)=C1 OFKRKCHCYWQZLY-XHBSWPGZSA-N 0.000 description 1
- 229950010594 cefsumide Drugs 0.000 description 1
- 229940036735 ceftaroline Drugs 0.000 description 1
- ZCCUWMICIWSJIX-NQJJCJBVSA-N ceftaroline fosamil Chemical compound S([C@@H]1[C@@H](C(N1C=1C([O-])=O)=O)NC(=O)\C(=N/OCC)C=2N=C(NP(O)(O)=O)SN=2)CC=1SC(SC=1)=NC=1C1=CC=[N+](C)C=C1 ZCCUWMICIWSJIX-NQJJCJBVSA-N 0.000 description 1
- 229960000484 ceftazidime Drugs 0.000 description 1
- ORFOPKXBNMVMKC-DWVKKRMSSA-N ceftazidime Chemical compound S([C@@H]1[C@@H](C(N1C=1C([O-])=O)=O)NC(=O)\C(=N/OC(C)(C)C(O)=O)C=2N=C(N)SC=2)CC=1C[N+]1=CC=CC=C1 ORFOPKXBNMVMKC-DWVKKRMSSA-N 0.000 description 1
- 229950000679 cefteram Drugs 0.000 description 1
- 229960004366 ceftezole Drugs 0.000 description 1
- DZMVCVMFETWNIU-LDYMZIIASA-N ceftezole Chemical compound O=C([C@@H](NC(=O)CN1N=NN=C1)[C@H]1SC2)N1C(C(=O)O)=C2CSC1=NN=CS1 DZMVCVMFETWNIU-LDYMZIIASA-N 0.000 description 1
- 229960004086 ceftibuten Drugs 0.000 description 1
- UNJFKXSSGBWRBZ-BJCIPQKHSA-N ceftibuten Chemical compound S1C(N)=NC(C(=C\CC(O)=O)\C(=O)N[C@@H]2C(N3C(=CCS[C@@H]32)C(O)=O)=O)=C1 UNJFKXSSGBWRBZ-BJCIPQKHSA-N 0.000 description 1
- 229960005229 ceftiofur Drugs 0.000 description 1
- ZBHXIWJRIFEVQY-IHMPYVIRSA-N ceftiofur Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC(=O)C1=CC=CO1 ZBHXIWJRIFEVQY-IHMPYVIRSA-N 0.000 description 1
- WJXAHFZIHLTPFR-JLRJEBFFSA-N ceftiolene Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1\C=C\SC1=NNC(=O)C(=O)N1CC=O WJXAHFZIHLTPFR-JLRJEBFFSA-N 0.000 description 1
- 229950008880 ceftiolene Drugs 0.000 description 1
- 229950007152 ceftioxide Drugs 0.000 description 1
- 229960001991 ceftizoxime Drugs 0.000 description 1
- NNULBSISHYWZJU-LLKWHZGFSA-N ceftizoxime Chemical compound N([C@@H]1C(N2C(=CCS[C@@H]21)C(O)=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 NNULBSISHYWZJU-LLKWHZGFSA-N 0.000 description 1
- VOAZJEPQLGBXGO-SDAWRPRTSA-N ceftobiprole Chemical compound S1C(N)=NC(C(=N\O)\C(=O)N[C@@H]2C(N3C(=C(\C=C/4C(N([C@H]5CNCC5)CC\4)=O)CS[C@@H]32)C(O)=O)=O)=N1 VOAZJEPQLGBXGO-SDAWRPRTSA-N 0.000 description 1
- 229950004259 ceftobiprole Drugs 0.000 description 1
- 229960002405 ceftolozane Drugs 0.000 description 1
- JHFNIHVVXRKLEF-DCZLAGFPSA-N ceftolozane Chemical compound CN1C(N)=C(NC(=O)NCCN)C=[N+]1CC1=C(C(O)=O)N2C(=O)[C@@H](NC(=O)C(=N/OC(C)(C)C([O-])=O)\C=3N=C(N)SN=3)[C@H]2SC1 JHFNIHVVXRKLEF-DCZLAGFPSA-N 0.000 description 1
- 229960004755 ceftriaxone Drugs 0.000 description 1
- VAAUVRVFOQPIGI-SPQHTLEESA-N ceftriaxone Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NC(=O)C(=O)NN1C VAAUVRVFOQPIGI-SPQHTLEESA-N 0.000 description 1
- 229950010799 cefuracetime Drugs 0.000 description 1
- 229960001668 cefuroxime Drugs 0.000 description 1
- JFPVXVDWJQMJEE-IZRZKJBUSA-N cefuroxime Chemical compound N([C@@H]1C(N2C(=C(COC(N)=O)CS[C@@H]21)C(O)=O)=O)C(=O)\C(=N/OC)C1=CC=CO1 JFPVXVDWJQMJEE-IZRZKJBUSA-N 0.000 description 1
- CXHKZHZLDMQGFF-ZSDSSEDPSA-N cefuzonam Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=CN=NS1 CXHKZHZLDMQGFF-ZSDSSEDPSA-N 0.000 description 1
- 229950000807 cefuzonam Drugs 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000006037 cell lysis Effects 0.000 description 1
- 238000001516 cell proliferation assay Methods 0.000 description 1
- 230000036755 cellular response Effects 0.000 description 1
- 229920001727 cellulose butyrate Polymers 0.000 description 1
- 229920006218 cellulose propionate Polymers 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 229940106164 cephalexin Drugs 0.000 description 1
- ZAIPMKNFIOOWCQ-UEKVPHQBSA-N cephalexin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)C)C(O)=O)=CC=CC=C1 ZAIPMKNFIOOWCQ-UEKVPHQBSA-N 0.000 description 1
- 229940124587 cephalosporin Drugs 0.000 description 1
- 150000001780 cephalosporins Chemical class 0.000 description 1
- VUFGUVLLDPOSBC-XRZFDKQNSA-M cephalothin sodium Chemical compound [Na+].N([C@H]1[C@@H]2N(C1=O)C(=C(CS2)COC(=O)C)C([O-])=O)C(=O)CC1=CC=CS1 VUFGUVLLDPOSBC-XRZFDKQNSA-M 0.000 description 1
- RDLPVSKMFDYCOR-UEKVPHQBSA-N cephradine Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)C(=C(CS3)C)C(O)=O)=CCC=CC1 RDLPVSKMFDYCOR-UEKVPHQBSA-N 0.000 description 1
- 229950010329 cethromycin Drugs 0.000 description 1
- 229940082500 cetostearyl alcohol Drugs 0.000 description 1
- 229960000541 cetyl alcohol Drugs 0.000 description 1
- CYETUYYEVKNSHZ-LGOOQLFJSA-N chembl1200498 Chemical compound CS(O)(=O)=O.C([C@@H]1[C@H]([C@@H]1C1)NC(=O)[C@H](C)NC(=O)[C@@H](N)C)N1C(C(=CC=1C(=O)C(C(O)=O)=C2)F)=NC=1N2C1=CC=C(F)C=C1F CYETUYYEVKNSHZ-LGOOQLFJSA-N 0.000 description 1
- DDTDNCYHLGRFBM-YZEKDTGTSA-N chembl2367892 Chemical compound CC(=O)N[C@H]1[C@@H](O)[C@H](O)[C@H](CO)O[C@H]1O[C@@H]([C@H]1C(N[C@@H](C2=CC(O)=CC(O[C@@H]3[C@H]([C@H](O)[C@H](O)[C@@H](CO)O3)O)=C2C=2C(O)=CC=C(C=2)[C@@H](NC(=O)[C@@H]2NC(=O)[C@@H]3C=4C=C(O)C=C(C=4)OC=4C(O)=CC=C(C=4)[C@@H](N)C(=O)N[C@H](CC=4C=C(Cl)C(O5)=CC=4)C(=O)N3)C(=O)N1)C(O)=O)=O)C(C=C1Cl)=CC=C1OC1=C(O[C@H]3[C@H]([C@@H](O)[C@H](O)[C@H](CO)O3)NC(C)=O)C5=CC2=C1 DDTDNCYHLGRFBM-YZEKDTGTSA-N 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- 210000000349 chromosome Anatomy 0.000 description 1
- 208000037976 chronic inflammation Diseases 0.000 description 1
- 208000037893 chronic inflammatory disorder Diseases 0.000 description 1
- DHSUYTOATWAVLW-WFVMDLQDSA-N cilastatin Chemical compound CC1(C)C[C@@H]1C(=O)N\C(=C/CCCCSC[C@H](N)C(O)=O)C(O)=O DHSUYTOATWAVLW-WFVMDLQDSA-N 0.000 description 1
- 229960004912 cilastatin Drugs 0.000 description 1
- 229960004621 cinoxacin Drugs 0.000 description 1
- VDUWPHTZYNWKRN-UHFFFAOYSA-N cinoxacin Chemical compound C1=C2N(CC)N=C(C(O)=O)C(=O)C2=CC2=C1OCO2 VDUWPHTZYNWKRN-UHFFFAOYSA-N 0.000 description 1
- 229960003405 ciprofloxacin Drugs 0.000 description 1
- 239000007979 citrate buffer Substances 0.000 description 1
- 235000015165 citric acid Nutrition 0.000 description 1
- 239000009194 citrus pectin Substances 0.000 description 1
- 229940040387 citrus pectin Drugs 0.000 description 1
- 229960002626 clarithromycin Drugs 0.000 description 1
- AGOYDEPGAOXOCK-KCBOHYOISA-N clarithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@](C)([C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)OC)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 AGOYDEPGAOXOCK-KCBOHYOISA-N 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 229950001320 clinafloxacin Drugs 0.000 description 1
- QGPKADBNRMWEQR-UHFFFAOYSA-N clinafloxacin Chemical compound C1C(N)CCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN(C3CC3)C2=C1Cl QGPKADBNRMWEQR-UHFFFAOYSA-N 0.000 description 1
- 229960004287 clofazimine Drugs 0.000 description 1
- WDQPAMHFFCXSNU-BGABXYSRSA-N clofazimine Chemical compound C12=CC=CC=C2N=C2C=C(NC=3C=CC(Cl)=CC=3)C(=N/C(C)C)/C=C2N1C1=CC=C(Cl)C=C1 WDQPAMHFFCXSNU-BGABXYSRSA-N 0.000 description 1
- 229960003326 cloxacillin Drugs 0.000 description 1
- LQOLIRLGBULYKD-JKIFEVAISA-N cloxacillin Chemical compound N([C@@H]1C(N2[C@H](C(C)(C)S[C@@H]21)C(O)=O)=O)C(=O)C1=C(C)ON=C1C1=CC=CC=C1Cl LQOLIRLGBULYKD-JKIFEVAISA-N 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 229940110456 cocoa butter Drugs 0.000 description 1
- 235000019868 cocoa butter Nutrition 0.000 description 1
- 235000019864 coconut oil Nutrition 0.000 description 1
- 239000003240 coconut oil Substances 0.000 description 1
- 210000004953 colonic tissue Anatomy 0.000 description 1
- 238000004891 communication Methods 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 238000007906 compression Methods 0.000 description 1
- 230000006835 compression Effects 0.000 description 1
- 238000002591 computed tomography Methods 0.000 description 1
- 238000004624 confocal microscopy Methods 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 101150055601 cops2 gene Proteins 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 208000029078 coronary artery disease Diseases 0.000 description 1
- 210000004351 coronary vessel Anatomy 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000009260 cross reactivity Effects 0.000 description 1
- 235000010947 crosslinked sodium carboxy methyl cellulose Nutrition 0.000 description 1
- 239000001767 crosslinked sodium carboxy methyl cellulose Substances 0.000 description 1
- DMSZORWOGDLWGN-UHFFFAOYSA-N ctk1a3526 Chemical compound NP(N)(N)=O DMSZORWOGDLWGN-UHFFFAOYSA-N 0.000 description 1
- 230000001351 cycling effect Effects 0.000 description 1
- 229960003077 cycloserine Drugs 0.000 description 1
- 229960002488 dalbavancin Drugs 0.000 description 1
- 108700009376 dalbavancin Proteins 0.000 description 1
- 229960002615 dalfopristin Drugs 0.000 description 1
- SUYRLXYYZQTJHF-VMBLUXKRSA-N dalfopristin Chemical compound O=C([C@@H]1N(C2=O)CC[C@H]1S(=O)(=O)CCN(CC)CC)O[C@H](C(C)C)[C@H](C)\C=C\C(=O)NC\C=C\C(\C)=C\[C@@H](O)CC(=O)CC1=NC2=CO1 SUYRLXYYZQTJHF-VMBLUXKRSA-N 0.000 description 1
- 108700028430 dalfopristin Proteins 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 229960000860 dapsone Drugs 0.000 description 1
- 229960005484 daptomycin Drugs 0.000 description 1
- DOAKLVKFURWEDJ-QCMAZARJSA-N daptomycin Chemical compound C([C@H]1C(=O)O[C@H](C)[C@@H](C(NCC(=O)N[C@@H](CCCN)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@H](CO)C(=O)N[C@H](C(=O)N1)[C@H](C)CC(O)=O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H](CC(N)=O)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)CCCCCCCCC)C(=O)C1=CC=CC=C1N DOAKLVKFURWEDJ-QCMAZARJSA-N 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- 229950006412 delafloxacin Drugs 0.000 description 1
- DYDCPNMLZGFQTM-UHFFFAOYSA-N delafloxacin Chemical compound C1=C(F)C(N)=NC(N2C3=C(Cl)C(N4CC(O)C4)=C(F)C=C3C(=O)C(C(O)=O)=C2)=C1F DYDCPNMLZGFQTM-UHFFFAOYSA-N 0.000 description 1
- 229960002398 demeclocycline Drugs 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 239000007933 dermal patch Substances 0.000 description 1
- 201000001981 dermatomyositis Diseases 0.000 description 1
- 239000003599 detergent Substances 0.000 description 1
- 235000019425 dextrin Nutrition 0.000 description 1
- 229960003807 dibekacin Drugs 0.000 description 1
- JJCQSGDBDPYCEO-XVZSLQNASA-N dibekacin Chemical compound O1[C@H](CN)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N JJCQSGDBDPYCEO-XVZSLQNASA-N 0.000 description 1
- PXBRQCKWGAHEHS-UHFFFAOYSA-N dichlorodifluoromethane Chemical compound FC(F)(Cl)Cl PXBRQCKWGAHEHS-UHFFFAOYSA-N 0.000 description 1
- 235000019404 dichlorodifluoromethane Nutrition 0.000 description 1
- 229940042935 dichlorodifluoromethane Drugs 0.000 description 1
- 229940087091 dichlorotetrafluoroethane Drugs 0.000 description 1
- 229960001585 dicloxacillin Drugs 0.000 description 1
- YFAGHNZHGGCZAX-JKIFEVAISA-N dicloxacillin Chemical compound N([C@@H]1C(N2[C@H](C(C)(C)S[C@@H]21)C(O)=O)=O)C(=O)C1=C(C)ON=C1C1=C(Cl)C=CC=C1Cl YFAGHNZHGGCZAX-JKIFEVAISA-N 0.000 description 1
- 235000013325 dietary fiber Nutrition 0.000 description 1
- 230000010339 dilation Effects 0.000 description 1
- 229940113088 dimethylacetamide Drugs 0.000 description 1
- 230000003292 diminished effect Effects 0.000 description 1
- NAGJZTKCGNOGPW-UHFFFAOYSA-K dioxido-sulfanylidene-sulfido-$l^{5}-phosphane Chemical compound [O-]P([O-])([S-])=S NAGJZTKCGNOGPW-UHFFFAOYSA-K 0.000 description 1
- 230000008034 disappearance Effects 0.000 description 1
- 230000009266 disease activity Effects 0.000 description 1
- 230000005750 disease progression Effects 0.000 description 1
- BFMYDTVEBKDAKJ-UHFFFAOYSA-L disodium;(2',7'-dibromo-3',6'-dioxido-3-oxospiro[2-benzofuran-1,9'-xanthene]-4'-yl)mercury;hydrate Chemical compound O.[Na+].[Na+].O1C(=O)C2=CC=CC=C2C21C1=CC(Br)=C([O-])C([Hg])=C1OC1=C2C=C(Br)C([O-])=C1 BFMYDTVEBKDAKJ-UHFFFAOYSA-L 0.000 description 1
- 108010007093 dispase Proteins 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- MYRTYDVEIRVNKP-UHFFFAOYSA-N divinylbenzene Substances C=CC1=CC=CC=C1C=C MYRTYDVEIRVNKP-UHFFFAOYSA-N 0.000 description 1
- 229960000895 doripenem Drugs 0.000 description 1
- AVAACINZEOAHHE-VFZPANTDSA-N doripenem Chemical compound C=1([C@H](C)[C@@H]2[C@H](C(N2C=1C(O)=O)=O)[C@H](O)C)S[C@@H]1CN[C@H](CNS(N)(=O)=O)C1 AVAACINZEOAHHE-VFZPANTDSA-N 0.000 description 1
- 229960003722 doxycycline Drugs 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 108010046161 drug combination polymyxin B neomycin sulfate bacitracin zinc Proteins 0.000 description 1
- 229940112141 dry powder inhaler Drugs 0.000 description 1
- 230000009977 dual effect Effects 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 229960002549 enoxacin Drugs 0.000 description 1
- IDYZIJYBMGIQMJ-UHFFFAOYSA-N enoxacin Chemical compound N1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC(F)=C1N1CCNCC1 IDYZIJYBMGIQMJ-UHFFFAOYSA-N 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- YQGOJNYOYNNSMM-UHFFFAOYSA-N eosin Chemical compound [Na+].OC(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C(O)=C(Br)C=C21 YQGOJNYOYNNSMM-UHFFFAOYSA-N 0.000 description 1
- 229960002457 epicillin Drugs 0.000 description 1
- RPBAFSBGYDKNRG-NJBDSQKTSA-N epicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CCC=CC1 RPBAFSBGYDKNRG-NJBDSQKTSA-N 0.000 description 1
- 210000005081 epithelial layer Anatomy 0.000 description 1
- 229960002770 ertapenem Drugs 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 229960000285 ethambutol Drugs 0.000 description 1
- AEOCXXJPGCBFJA-UHFFFAOYSA-N ethionamide Chemical compound CCC1=CC(C(N)=S)=CC=N1 AEOCXXJPGCBFJA-UHFFFAOYSA-N 0.000 description 1
- 229960002001 ethionamide Drugs 0.000 description 1
- LVGKNOAMLMIIKO-QXMHVHEDSA-N ethyl oleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OCC LVGKNOAMLMIIKO-QXMHVHEDSA-N 0.000 description 1
- 229940093471 ethyl oleate Drugs 0.000 description 1
- 238000013265 extended release Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 235000013861 fat-free Nutrition 0.000 description 1
- 150000002191 fatty alcohols Chemical class 0.000 description 1
- 230000004761 fibrosis Effects 0.000 description 1
- 230000003176 fibrotic effect Effects 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 229960003306 fleroxacin Drugs 0.000 description 1
- XBJBPGROQZJDOJ-UHFFFAOYSA-N fleroxacin Chemical compound C1CN(C)CCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN(CCF)C2=C1F XBJBPGROQZJDOJ-UHFFFAOYSA-N 0.000 description 1
- 229960002878 flomoxef Drugs 0.000 description 1
- UHRBTBZOWWGKMK-DOMZBBRYSA-N flomoxef Chemical compound O([C@@H]1[C@@](C(N1C=1C(O)=O)=O)(NC(=O)CSC(F)F)OC)CC=1CSC1=NN=NN1CCO UHRBTBZOWWGKMK-DOMZBBRYSA-N 0.000 description 1
- 238000001943 fluorescence-activated cell sorting Methods 0.000 description 1
- 229960000308 fosfomycin Drugs 0.000 description 1
- YMDXZJFXQJVXBF-STHAYSLISA-N fosfomycin Chemical compound C[C@@H]1O[C@@H]1P(O)(O)=O YMDXZJFXQJVXBF-STHAYSLISA-N 0.000 description 1
- 231100000221 frame shift mutation induction Toxicity 0.000 description 1
- 230000037433 frameshift Effects 0.000 description 1
- FTSSQIKWUOOEGC-RULYVFMPSA-N fructooligosaccharide Chemical compound OC[C@H]1O[C@@](CO)(OC[C@@]2(OC[C@@]3(OC[C@@]4(OC[C@@]5(OC[C@@]6(OC[C@@]7(OC[C@@]8(OC[C@@]9(OC[C@@]%10(OC[C@@]%11(O[C@H]%12O[C@H](CO)[C@@H](O)[C@H](O)[C@H]%12O)O[C@H](CO)[C@@H](O)[C@@H]%11O)O[C@H](CO)[C@@H](O)[C@@H]%10O)O[C@H](CO)[C@@H](O)[C@@H]9O)O[C@H](CO)[C@@H](O)[C@@H]8O)O[C@H](CO)[C@@H](O)[C@@H]7O)O[C@H](CO)[C@@H](O)[C@@H]6O)O[C@H](CO)[C@@H](O)[C@@H]5O)O[C@H](CO)[C@@H](O)[C@@H]4O)O[C@H](CO)[C@@H](O)[C@@H]3O)O[C@H](CO)[C@@H](O)[C@@H]2O)[C@@H](O)[C@@H]1O FTSSQIKWUOOEGC-RULYVFMPSA-N 0.000 description 1
- 229940107187 fructooligosaccharide Drugs 0.000 description 1
- 239000001530 fumaric acid Substances 0.000 description 1
- 229960001625 furazolidone Drugs 0.000 description 1
- PLHJDBGFXBMTGZ-WEVVVXLNSA-N furazolidone Chemical compound O1C([N+](=O)[O-])=CC=C1\C=N\N1C(=O)OCC1 PLHJDBGFXBMTGZ-WEVVVXLNSA-N 0.000 description 1
- 229960004675 fusidic acid Drugs 0.000 description 1
- IECPWNUMDGFDKC-MZJAQBGESA-N fusidic acid Chemical compound O[C@@H]([C@@H]12)C[C@H]3\C(=C(/CCC=C(C)C)C(O)=O)[C@@H](OC(C)=O)C[C@]3(C)[C@@]2(C)CC[C@@H]2[C@]1(C)CC[C@@H](O)[C@H]2C IECPWNUMDGFDKC-MZJAQBGESA-N 0.000 description 1
- 235000021255 galacto-oligosaccharides Nutrition 0.000 description 1
- 150000003271 galactooligosaccharides Chemical class 0.000 description 1
- 229960003923 gatifloxacin Drugs 0.000 description 1
- 238000003304 gavage Methods 0.000 description 1
- QTQAWLPCGQOSGP-GBTDJJJQSA-N geldanamycin Chemical compound N1C(=O)\C(C)=C/C=C\[C@@H](OC)[C@H](OC(N)=O)\C(C)=C/[C@@H](C)[C@@H](O)[C@H](OC)C[C@@H](C)CC2=C(OC)C(=O)C=C1C2=O QTQAWLPCGQOSGP-GBTDJJJQSA-N 0.000 description 1
- 229960003170 gemifloxacin Drugs 0.000 description 1
- ZRCVYEYHRGVLOC-HYARGMPZSA-N gemifloxacin Chemical compound C1C(CN)C(=N/OC)/CN1C(C(=C1)F)=NC2=C1C(=O)C(C(O)=O)=CN2C1CC1 ZRCVYEYHRGVLOC-HYARGMPZSA-N 0.000 description 1
- 238000010363 gene targeting Methods 0.000 description 1
- 238000003205 genotyping method Methods 0.000 description 1
- 208000018090 giant cell myocarditis Diseases 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 229940046240 glucomannan Drugs 0.000 description 1
- 239000000174 gluconic acid Substances 0.000 description 1
- 235000012208 gluconic acid Nutrition 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 229940049654 glyceryl behenate Drugs 0.000 description 1
- 150000002334 glycols Chemical class 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 229960000642 grepafloxacin Drugs 0.000 description 1
- 230000036433 growing body Effects 0.000 description 1
- 239000000665 guar gum Substances 0.000 description 1
- 235000010417 guar gum Nutrition 0.000 description 1
- 229960002154 guar gum Drugs 0.000 description 1
- 150000008282 halocarbons Chemical class 0.000 description 1
- IUJAMGNYPWYUPM-UHFFFAOYSA-N hentriacontane Chemical compound CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCC IUJAMGNYPWYUPM-UHFFFAOYSA-N 0.000 description 1
- MCAHMSDENAOJFZ-BVXDHVRPSA-N herbimycin Chemical compound N1C(=O)\C(C)=C\C=C/[C@H](OC)[C@@H](OC(N)=O)\C(C)=C\[C@H](C)[C@@H](OC)[C@@H](OC)C[C@H](C)[C@@H](OC)C2=CC(=O)C=C1C2=O MCAHMSDENAOJFZ-BVXDHVRPSA-N 0.000 description 1
- 229930193320 herbimycin Natural products 0.000 description 1
- 229960003884 hetacillin Drugs 0.000 description 1
- DXVUYOAEDJXBPY-NFFDBFGFSA-N hetacillin Chemical compound C1([C@@H]2C(=O)N(C(N2)(C)C)[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 DXVUYOAEDJXBPY-NFFDBFGFSA-N 0.000 description 1
- 238000007489 histopathology method Methods 0.000 description 1
- 230000013632 homeostatic process Effects 0.000 description 1
- 244000005702 human microbiome Species 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 150000005828 hydrofluoroalkanes Chemical class 0.000 description 1
- 239000000017 hydrogel Substances 0.000 description 1
- 235000019447 hydroxyethyl cellulose Nutrition 0.000 description 1
- 229920003063 hydroxymethyl cellulose Polymers 0.000 description 1
- 229940031574 hydroxymethyl cellulose Drugs 0.000 description 1
- 229960002182 imipenem Drugs 0.000 description 1
- ZSKVGTPCRGIANV-ZXFLCMHBSA-N imipenem Chemical compound C1C(SCC\N=C\N)=C(C(O)=O)N2C(=O)[C@H]([C@H](O)C)[C@H]21 ZSKVGTPCRGIANV-ZXFLCMHBSA-N 0.000 description 1
- 230000005934 immune activation Effects 0.000 description 1
- 230000005746 immune checkpoint blockade Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 238000010166 immunofluorescence Methods 0.000 description 1
- 238000000530 impalefection Methods 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000012405 in silico analysis Methods 0.000 description 1
- 238000000126 in silico method Methods 0.000 description 1
- 230000002458 infectious effect Effects 0.000 description 1
- 230000008595 infiltration Effects 0.000 description 1
- 238000001764 infiltration Methods 0.000 description 1
- 210000004968 inflammatory monocyte/macrophage Anatomy 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 108010043603 integrin alpha4beta7 Proteins 0.000 description 1
- 210000005206 intestinal lamina propria Anatomy 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- PGHMRUGBZOYCAA-UHFFFAOYSA-N ionomycin Natural products O1C(CC(O)C(C)C(O)C(C)C=CCC(C)CC(C)C(O)=CC(=O)C(C)CC(C)CC(CCC(O)=O)C)CCC1(C)C1OC(C)(C(C)O)CC1 PGHMRUGBZOYCAA-UHFFFAOYSA-N 0.000 description 1
- PGHMRUGBZOYCAA-ADZNBVRBSA-N ionomycin Chemical compound O1[C@H](C[C@H](O)[C@H](C)[C@H](O)[C@H](C)/C=C/C[C@@H](C)C[C@@H](C)C(/O)=C/C(=O)[C@@H](C)C[C@@H](C)C[C@@H](CCC(O)=O)C)CC[C@@]1(C)[C@@H]1O[C@](C)([C@@H](C)O)CC1 PGHMRUGBZOYCAA-ADZNBVRBSA-N 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 229960003350 isoniazid Drugs 0.000 description 1
- QRXWMOHMRWLFEY-UHFFFAOYSA-N isoniazide Chemical compound NNC(=O)C1=CC=NC=C1 QRXWMOHMRWLFEY-UHFFFAOYSA-N 0.000 description 1
- 229960004144 josamycin Drugs 0.000 description 1
- XJSFLOJWULLJQS-NGVXBBESSA-N josamycin Chemical compound CO[C@H]1[C@H](OC(C)=O)CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@H]1[C@H](O)[C@@H](N(C)C)[C@H](O[C@@H]2O[C@@H](C)[C@H](OC(=O)CC(C)C)[C@](C)(O)C2)[C@@H](C)O1 XJSFLOJWULLJQS-NGVXBBESSA-N 0.000 description 1
- 229960000318 kanamycin Drugs 0.000 description 1
- 229930027917 kanamycin Natural products 0.000 description 1
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 1
- 229930182823 kanamycin A Natural products 0.000 description 1
- BJHIKXHVCXFQLS-PQLUHFTBSA-N keto-D-tagatose Chemical compound OC[C@@H](O)[C@H](O)[C@H](O)C(=O)CO BJHIKXHVCXFQLS-PQLUHFTBSA-N 0.000 description 1
- 239000003835 ketolide antibiotic agent Substances 0.000 description 1
- 229950007634 kitasamycin Drugs 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- JCQLYHFGKNRPGE-FCVZTGTOSA-N lactulose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 JCQLYHFGKNRPGE-FCVZTGTOSA-N 0.000 description 1
- 229960000511 lactulose Drugs 0.000 description 1
- PFCRQPBOOFTZGQ-UHFFFAOYSA-N lactulose keto form Natural products OCC(=O)C(O)C(C(O)CO)OC1OC(CO)C(O)C(O)C1O PFCRQPBOOFTZGQ-UHFFFAOYSA-N 0.000 description 1
- 229940039717 lanolin Drugs 0.000 description 1
- 235000019388 lanolin Nutrition 0.000 description 1
- 229960000433 latamoxef Drugs 0.000 description 1
- 210000005240 left ventricle Anatomy 0.000 description 1
- XYJOGTQLTFNMQG-KJHBSLKPSA-N leucomycin V Chemical compound CO[C@H]1[C@H](O)CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@H]1[C@H](O)[C@@H](N(C)C)[C@H](O[C@@H]2O[C@@H](C)[C@H](O)[C@](C)(O)C2)[C@@H](C)O1 XYJOGTQLTFNMQG-KJHBSLKPSA-N 0.000 description 1
- 229960003376 levofloxacin Drugs 0.000 description 1
- 229960003907 linezolid Drugs 0.000 description 1
- TYZROVQLWOKYKF-ZDUSSCGKSA-N linezolid Chemical compound O=C1O[C@@H](CNC(=O)C)CN1C(C=C1F)=CC=C1N1CCOCC1 TYZROVQLWOKYKF-ZDUSSCGKSA-N 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 230000004807 localization Effects 0.000 description 1
- 229960002422 lomefloxacin Drugs 0.000 description 1
- ZEKZLJVOYLTDKK-UHFFFAOYSA-N lomefloxacin Chemical compound FC1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC(F)=C1N1CCNC(C)C1 ZEKZLJVOYLTDKK-UHFFFAOYSA-N 0.000 description 1
- 239000007937 lozenge Substances 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 210000005210 lymphoid organ Anatomy 0.000 description 1
- 239000012139 lysis buffer Substances 0.000 description 1
- 239000003120 macrolide antibiotic agent Substances 0.000 description 1
- 229940041033 macrolides Drugs 0.000 description 1
- VZCYOOQTPOCHFL-UPHRSURJSA-N maleic acid Chemical compound OC(=O)\C=C/C(O)=O VZCYOOQTPOCHFL-UPHRSURJSA-N 0.000 description 1
- 239000011976 maleic acid Substances 0.000 description 1
- 210000001161 mammalian embryo Anatomy 0.000 description 1
- 150000003272 mannan oligosaccharides Chemical class 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- 229960002260 meropenem Drugs 0.000 description 1
- DMJNNHOOLUXYBV-PQTSNVLCSA-N meropenem Chemical compound C=1([C@H](C)[C@@H]2[C@H](C(N2C=1C(O)=O)=O)[C@H](O)C)S[C@@H]1CN[C@H](C(=O)N(C)C)C1 DMJNNHOOLUXYBV-PQTSNVLCSA-N 0.000 description 1
- 229960003806 metampicillin Drugs 0.000 description 1
- FZECHKJQHUVANE-MCYUEQNJSA-N metampicillin Chemical compound C1([C@@H](N=C)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 FZECHKJQHUVANE-MCYUEQNJSA-N 0.000 description 1
- 125000005397 methacrylic acid ester group Chemical group 0.000 description 1
- 229960003085 meticillin Drugs 0.000 description 1
- 229960000198 mezlocillin Drugs 0.000 description 1
- YPBATNHYBCGSSN-VWPFQQQWSA-N mezlocillin Chemical compound N([C@@H](C(=O)N[C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C=1C=CC=CC=1)C(=O)N1CCN(S(C)(=O)=O)C1=O YPBATNHYBCGSSN-VWPFQQQWSA-N 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 229960002757 midecamycin Drugs 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 229940042472 mineral oil Drugs 0.000 description 1
- 150000007522 mineralic acids Chemical class 0.000 description 1
- 229960004023 minocycline Drugs 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 102000035118 modified proteins Human genes 0.000 description 1
- 108091005573 modified proteins Proteins 0.000 description 1
- 239000003068 molecular probe Substances 0.000 description 1
- 229940041009 monobactams Drugs 0.000 description 1
- 210000005087 mononuclear cell Anatomy 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- HWPKGOGLCKPRLZ-UHFFFAOYSA-M monosodium citrate Chemical compound [Na+].OC(=O)CC(O)(C([O-])=O)CC(O)=O HWPKGOGLCKPRLZ-UHFFFAOYSA-M 0.000 description 1
- 239000002524 monosodium citrate Substances 0.000 description 1
- 235000018342 monosodium citrate Nutrition 0.000 description 1
- 230000001002 morphogenetic effect Effects 0.000 description 1
- 229960003702 moxifloxacin Drugs 0.000 description 1
- FABPRXSRWADJSP-MEDUHNTESA-N moxifloxacin Chemical compound COC1=C(N2C[C@H]3NCCC[C@H]3C2)C(F)=CC(C(C(C(O)=O)=C2)=O)=C1N2C1CC1 FABPRXSRWADJSP-MEDUHNTESA-N 0.000 description 1
- 229960003128 mupirocin Drugs 0.000 description 1
- 229930187697 mupirocin Natural products 0.000 description 1
- DDHVILIIHBIMQU-YJGQQKNPSA-L mupirocin calcium hydrate Chemical compound O.O.[Ca+2].C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC([O-])=O)OC1.C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC([O-])=O)OC1 DDHVILIIHBIMQU-YJGQQKNPSA-L 0.000 description 1
- 208000010125 myocardial infarction Diseases 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- JORAUNFTUVJTNG-BSTBCYLQSA-N n-[(2s)-4-amino-1-[[(2s,3r)-1-[[(2s)-4-amino-1-oxo-1-[[(3s,6s,9s,12s,15r,18s,21s)-6,9,18-tris(2-aminoethyl)-3-[(1r)-1-hydroxyethyl]-12,15-bis(2-methylpropyl)-2,5,8,11,14,17,20-heptaoxo-1,4,7,10,13,16,19-heptazacyclotricos-21-yl]amino]butan-2-yl]amino]-3-h Polymers CC(C)CCCCC(=O)N[C@@H](CCN)C(=O)N[C@H]([C@@H](C)O)CN[C@@H](CCN)C(=O)N[C@H]1CCNC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCN)NC(=O)[C@H](CCN)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CCN)NC1=O.CCC(C)CCCCC(=O)N[C@@H](CCN)C(=O)N[C@H]([C@@H](C)O)CN[C@@H](CCN)C(=O)N[C@H]1CCNC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCN)NC(=O)[C@H](CCN)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CCN)NC1=O JORAUNFTUVJTNG-BSTBCYLQSA-N 0.000 description 1
- YKQOSKADJPQZHB-YNWHQGOSSA-N n-[(2s)-4-amino-1-[[(2s,3r)-1-[[(2s)-4-amino-1-oxo-1-[[(3s,6s,9s,12s,15r,18s,21s)-6,9,18-tris(2-aminoethyl)-3-[(1s)-1-hydroxyethyl]-12,15-bis(2-methylpropyl)-2,5,8,11,14,17,20-heptaoxo-1,4,7,10,13,16,19-heptazacyclotricos-21-yl]amino]butan-2-yl]amino]-3-h Polymers CCC(C)CCCC(=O)N[C@@H](CCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCN)C(=O)N[C@H]1CCNC(=O)[C@H]([C@H](C)O)NC(=O)[C@H](CCN)NC(=O)[C@H](CCN)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CCN)NC1=O YKQOSKADJPQZHB-YNWHQGOSSA-N 0.000 description 1
- GOQYKNQRPGWPLP-UHFFFAOYSA-N n-heptadecyl alcohol Natural products CCCCCCCCCCCCCCCCCO GOQYKNQRPGWPLP-UHFFFAOYSA-N 0.000 description 1
- 229960003808 nadifloxacin Drugs 0.000 description 1
- JYJTVFIEFKZWCJ-UHFFFAOYSA-N nadifloxacin Chemical compound FC1=CC(C(C(C(O)=O)=C2)=O)=C3N2C(C)CCC3=C1N1CCC(O)CC1 JYJTVFIEFKZWCJ-UHFFFAOYSA-N 0.000 description 1
- 229960000515 nafcillin Drugs 0.000 description 1
- GPXLMGHLHQJAGZ-JTDSTZFVSA-N nafcillin Chemical compound C1=CC=CC2=C(C(=O)N[C@@H]3C(N4[C@H](C(C)(C)S[C@@H]43)C(O)=O)=O)C(OCC)=CC=C21 GPXLMGHLHQJAGZ-JTDSTZFVSA-N 0.000 description 1
- 239000002105 nanoparticle Substances 0.000 description 1
- 239000007922 nasal spray Substances 0.000 description 1
- 229960002353 nemonoxacin Drugs 0.000 description 1
- AVPQPGFLVZTJOR-RYUDHWBXSA-N nemonoxacin Chemical compound COC1=C(N2C[C@@H](N)C[C@H](C)C2)C=CC(C(C(C(O)=O)=C2)=O)=C1N2C1CC1 AVPQPGFLVZTJOR-RYUDHWBXSA-N 0.000 description 1
- 229960004927 neomycin Drugs 0.000 description 1
- 229940049337 neosporin Drugs 0.000 description 1
- 229960000808 netilmicin Drugs 0.000 description 1
- ZBGPYVZLYBDXKO-HILBYHGXSA-N netilmycin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@]([C@H](NC)[C@@H](O)CO1)(C)O)NCC)[C@H]1OC(CN)=CC[C@H]1N ZBGPYVZLYBDXKO-HILBYHGXSA-N 0.000 description 1
- 238000007481 next generation sequencing Methods 0.000 description 1
- 229960000564 nitrofurantoin Drugs 0.000 description 1
- NXFQHRVNIOXGAQ-YCRREMRBSA-N nitrofurantoin Chemical compound O1C([N+](=O)[O-])=CC=C1\C=N\N1C(=O)NC(=O)C1 NXFQHRVNIOXGAQ-YCRREMRBSA-N 0.000 description 1
- 230000009871 nonspecific binding Effects 0.000 description 1
- 229960001180 norfloxacin Drugs 0.000 description 1
- OGJPXUAPXNRGGI-UHFFFAOYSA-N norfloxacin Chemical compound C1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC(F)=C1N1CCNCC1 OGJPXUAPXNRGGI-UHFFFAOYSA-N 0.000 description 1
- 230000001473 noxious effect Effects 0.000 description 1
- 238000010899 nucleation Methods 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 230000030648 nucleus localization Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000019895 oat fiber Nutrition 0.000 description 1
- 229960001699 ofloxacin Drugs 0.000 description 1
- 229960002351 oleandomycin Drugs 0.000 description 1
- 235000019367 oleandomycin Nutrition 0.000 description 1
- RZPAKFUAFGMUPI-KGIGTXTPSA-N oleandomycin Chemical compound O1[C@@H](C)[C@H](O)[C@@H](OC)C[C@@H]1O[C@@H]1[C@@H](C)C(=O)O[C@H](C)[C@H](C)[C@H](O)[C@@H](C)C(=O)[C@]2(OC2)C[C@H](C)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C RZPAKFUAFGMUPI-KGIGTXTPSA-N 0.000 description 1
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical class CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 229960001607 oritavancin Drugs 0.000 description 1
- 108010006945 oritavancin Proteins 0.000 description 1
- VHFGEBVPHAGQPI-MYYQHNLBSA-N oritavancin Chemical compound O([C@@H]1C2=CC=C(C(=C2)Cl)OC=2C=C3C=C(C=2O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)O[C@@H]2O[C@@H](C)[C@H](O)[C@@](C)(NCC=4C=CC(=CC=4)C=4C=CC(Cl)=CC=4)C2)OC2=CC=C(C=C2Cl)[C@@H](O)[C@H](C(N[C@@H](CC(N)=O)C(=O)N[C@H]3C(=O)N[C@H]2C(=O)N[C@@H]1C(N[C@H](C1=CC(O)=CC(O)=C1C=1C(O)=CC=C2C=1)C(O)=O)=O)=O)NC(=O)[C@@H](CC(C)C)NC)[C@H]1C[C@](C)(N)[C@@H](O)[C@H](C)O1 VHFGEBVPHAGQPI-MYYQHNLBSA-N 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- GSSMIHQEWAQUPM-AOLPDKKJSA-N ovalbumin peptide Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](N)[C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1NC=NC=1)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C1=CN=CN1 GSSMIHQEWAQUPM-AOLPDKKJSA-N 0.000 description 1
- 229960001019 oxacillin Drugs 0.000 description 1
- UWYHMGVUTGAWSP-JKIFEVAISA-N oxacillin Chemical compound N([C@@H]1C(N2[C@H](C(C)(C)S[C@@H]21)C(O)=O)=O)C(=O)C1=C(C)ON=C1C1=CC=CC=C1 UWYHMGVUTGAWSP-JKIFEVAISA-N 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 229960000321 oxolinic acid Drugs 0.000 description 1
- 229960000625 oxytetracycline Drugs 0.000 description 1
- IWVCMVBTMGNXQD-PXOLEDIWSA-N oxytetracycline Chemical compound C1=CC=C2[C@](O)(C)[C@H]3[C@H](O)[C@H]4[C@H](N(C)C)C(O)=C(C(N)=O)C(=O)[C@@]4(O)C(O)=C3C(=O)C2=C1O IWVCMVBTMGNXQD-PXOLEDIWSA-N 0.000 description 1
- 235000019366 oxytetracycline Nutrition 0.000 description 1
- LSQZJLSUYDQPKJ-UHFFFAOYSA-N p-Hydroxyampicillin Natural products O=C1N2C(C(O)=O)C(C)(C)SC2C1NC(=O)C(N)C1=CC=C(O)C=C1 LSQZJLSUYDQPKJ-UHFFFAOYSA-N 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- FJKROLUGYXJWQN-UHFFFAOYSA-N papa-hydroxy-benzoic acid Natural products OC(=O)C1=CC=C(O)C=C1 FJKROLUGYXJWQN-UHFFFAOYSA-N 0.000 description 1
- 235000019809 paraffin wax Nutrition 0.000 description 1
- 229920002866 paraformaldehyde Polymers 0.000 description 1
- 229960001914 paromomycin Drugs 0.000 description 1
- UOZODPSAJZTQNH-LSWIJEOBSA-N paromomycin Chemical compound N[C@@H]1[C@@H](O)[C@H](O)[C@H](CN)O[C@@H]1O[C@H]1[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](N)C[C@@H](N)[C@@H]2O)O[C@@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)O[C@@H]1CO UOZODPSAJZTQNH-LSWIJEOBSA-N 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 229960002625 pazufloxacin Drugs 0.000 description 1
- 229960004236 pefloxacin Drugs 0.000 description 1
- FHFYDNQZQSQIAI-UHFFFAOYSA-N pefloxacin Chemical compound C1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC(F)=C1N1CCN(C)CC1 FHFYDNQZQSQIAI-UHFFFAOYSA-N 0.000 description 1
- 239000004466 pelleted feed Substances 0.000 description 1
- 235000019371 penicillin G benzathine Nutrition 0.000 description 1
- MIFYHUACUWQUKT-GPUHXXMPSA-N penicillin N Chemical compound OC(=O)[C@H]1C(C)(C)S[C@@H]2[C@H](NC(=O)CCC[C@@H](N)C(O)=O)C(=O)N21 MIFYHUACUWQUKT-GPUHXXMPSA-N 0.000 description 1
- QULKGELYPOJSLP-WCABBAIRSA-N penicillin O Chemical compound OC(=O)[C@H]1C(C)(C)S[C@@H]2[C@H](NC(=O)CSCC=C)C(=O)N21 QULKGELYPOJSLP-WCABBAIRSA-N 0.000 description 1
- 229940056367 penicillin v Drugs 0.000 description 1
- 150000002960 penicillins Chemical class 0.000 description 1
- 230000008823 permeabilization Effects 0.000 description 1
- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 239000008194 pharmaceutical composition Substances 0.000 description 1
- BPLBGHOLXOTWMN-MBNYWOFBSA-N phenoxymethylpenicillin Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)COC1=CC=CC=C1 BPLBGHOLXOTWMN-MBNYWOFBSA-N 0.000 description 1
- 239000006187 pill Substances 0.000 description 1
- 229960001732 pipemidic acid Drugs 0.000 description 1
- JOHZPMXAZQZXHR-UHFFFAOYSA-N pipemidic acid Chemical compound N1=C2N(CC)C=C(C(O)=O)C(=O)C2=CN=C1N1CCNCC1 JOHZPMXAZQZXHR-UHFFFAOYSA-N 0.000 description 1
- 229960002292 piperacillin Drugs 0.000 description 1
- WCMIIGXFCMNQDS-IDYPWDAWSA-M piperacillin sodium Chemical compound [Na+].O=C1C(=O)N(CC)CCN1C(=O)N[C@H](C=1C=CC=CC=1)C(=O)N[C@@H]1C(=O)N2[C@@H](C([O-])=O)C(C)(C)S[C@@H]21 WCMIIGXFCMNQDS-IDYPWDAWSA-M 0.000 description 1
- 229960001635 pirlimycin Drugs 0.000 description 1
- 229960004444 piromidic acid Drugs 0.000 description 1
- RCIMBBZXSXFZBV-UHFFFAOYSA-N piromidic acid Chemical compound N1=C2N(CC)C=C(C(O)=O)C(=O)C2=CN=C1N1CCCC1 RCIMBBZXSXFZBV-UHFFFAOYSA-N 0.000 description 1
- 229960003342 pivampicillin Drugs 0.000 description 1
- ZEMIJUDPLILVNQ-ZXFNITATSA-N pivampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)[C@H](C(S3)(C)C)C(=O)OCOC(=O)C(C)(C)C)=CC=CC=C1 ZEMIJUDPLILVNQ-ZXFNITATSA-N 0.000 description 1
- 239000013600 plasmid vector Substances 0.000 description 1
- CSOMAHTTWTVBFL-OFBLZTNGSA-N platensimycin Chemical compound C([C@]1([C@@H]2[C@@H]3C[C@@H]4C[C@@]2(C=CC1=O)C[C@@]4(O3)C)C)CC(=O)NC1=C(O)C=CC(C(O)=O)=C1O CSOMAHTTWTVBFL-OFBLZTNGSA-N 0.000 description 1
- CSOMAHTTWTVBFL-UHFFFAOYSA-N platensimycin Natural products O1C2(C)CC3(C=CC4=O)CC2CC1C3C4(C)CCC(=O)NC1=C(O)C=CC(C(O)=O)=C1O CSOMAHTTWTVBFL-UHFFFAOYSA-N 0.000 description 1
- 229920000747 poly(lactic acid) Polymers 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000058 polyacrylate Polymers 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000004633 polyglycolic acid Substances 0.000 description 1
- 239000004626 polylactic acid Substances 0.000 description 1
- 102000054765 polymorphisms of proteins Human genes 0.000 description 1
- 229920000024 polymyxin B Polymers 0.000 description 1
- 108010046630 polymyxin B drug combination bacitracin Proteins 0.000 description 1
- XDJYMJULXQKGMM-UHFFFAOYSA-N polymyxin E1 Natural products CCC(C)CCCCC(=O)NC(CCN)C(=O)NC(C(C)O)C(=O)NC(CCN)C(=O)NC1CCNC(=O)C(C(C)O)NC(=O)C(CCN)NC(=O)C(CCN)NC(=O)C(CC(C)C)NC(=O)C(CC(C)C)NC(=O)C(CCN)NC1=O XDJYMJULXQKGMM-UHFFFAOYSA-N 0.000 description 1
- 229960005266 polymyxin b Drugs 0.000 description 1
- 229940041153 polymyxins Drugs 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 239000000244 polyoxyethylene sorbitan monooleate Substances 0.000 description 1
- 229940068968 polysorbate 80 Drugs 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 229940103255 polysporin Drugs 0.000 description 1
- 229920002223 polystyrene Polymers 0.000 description 1
- 239000011118 polyvinyl acetate Substances 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 229950004447 posizolid Drugs 0.000 description 1
- LLLCSBYSPJHDJX-UHFFFAOYSA-M potassium;2-methylprop-2-enoate Chemical compound [K+].CC(=C)C([O-])=O LLLCSBYSPJHDJX-UHFFFAOYSA-M 0.000 description 1
- 244000144977 poultry Species 0.000 description 1
- 229920003124 powdered cellulose Polymers 0.000 description 1
- 235000019814 powdered cellulose Nutrition 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000000513 principal component analysis Methods 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 108060006633 protein kinase Proteins 0.000 description 1
- 229960001224 prulifloxacin Drugs 0.000 description 1
- 229960005206 pyrazinamide Drugs 0.000 description 1
- IPEHBUMCGVEMRF-UHFFFAOYSA-N pyrazinecarboxamide Chemical compound NC(=O)C1=CN=CC=N1 IPEHBUMCGVEMRF-UHFFFAOYSA-N 0.000 description 1
- 239000001397 quillaja saponaria molina bark Substances 0.000 description 1
- 150000007660 quinolones Chemical class 0.000 description 1
- 229960005442 quinupristin Drugs 0.000 description 1
- WTHRRGMBUAHGNI-LCYNINFDSA-N quinupristin Chemical compound N([C@@H]1C(=O)N[C@@H](C(N2CCC[C@H]2C(=O)N(C)[C@@H](CC=2C=CC(=CC=2)N(C)C)C(=O)N2C[C@@H](CS[C@H]3C4CCN(CC4)C3)C(=O)C[C@H]2C(=O)N[C@H](C(=O)O[C@@H]1C)C=1C=CC=CC=1)=O)CC)C(=O)C1=NC=CC=C1O WTHRRGMBUAHGNI-LCYNINFDSA-N 0.000 description 1
- 108700028429 quinupristin Proteins 0.000 description 1
- 108700022487 rRNA Genes Proteins 0.000 description 1
- 229950009965 radezolid Drugs 0.000 description 1
- BTTNOGHPGJANSW-IBGZPJMESA-N radezolid Chemical compound O=C1O[C@@H](CNC(=O)C)CN1C1=CC=C(C=2C=CC(CNCC=3NN=NC=3)=CC=2)C(F)=C1 BTTNOGHPGJANSW-IBGZPJMESA-N 0.000 description 1
- 238000003753 real-time PCR Methods 0.000 description 1
- 239000006215 rectal suppository Substances 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000007634 remodeling Methods 0.000 description 1
- 230000008263 repair mechanism Effects 0.000 description 1
- 230000003362 replicative effect Effects 0.000 description 1
- 235000021254 resistant starch Nutrition 0.000 description 1
- 238000010839 reverse transcription Methods 0.000 description 1
- 239000009754 rhamnogalacturonan I Substances 0.000 description 1
- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 239000002336 ribonucleotide Substances 0.000 description 1
- 125000002652 ribonucleotide group Chemical group 0.000 description 1
- 229960000885 rifabutin Drugs 0.000 description 1
- 229960002599 rifapentine Drugs 0.000 description 1
- WDZCUPBHRAEYDL-GZAUEHORSA-N rifapentine Chemical compound O([C@](C1=O)(C)O/C=C/[C@@H]([C@H]([C@@H](OC(C)=O)[C@H](C)[C@H](O)[C@H](C)[C@@H](O)[C@@H](C)\C=C\C=C(C)/C(=O)NC=2C(O)=C3C(O)=C4C)C)OC)C4=C1C3=C(O)C=2\C=N\N(CC1)CCN1C1CCCC1 WDZCUPBHRAEYDL-GZAUEHORSA-N 0.000 description 1
- 229960003889 rosoxacin Drugs 0.000 description 1
- XBPZXDSZHPDXQU-UHFFFAOYSA-N rosoxacin Chemical compound C1=C2N(CC)C=C(C(O)=O)C(=O)C2=CC=C1C1=CC=NC=C1 XBPZXDSZHPDXQU-UHFFFAOYSA-N 0.000 description 1
- 229960005224 roxithromycin Drugs 0.000 description 1
- 229960004062 rufloxacin Drugs 0.000 description 1
- 229960004889 salicylic acid Drugs 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000007480 sanger sequencing Methods 0.000 description 1
- 229930182490 saponin Natural products 0.000 description 1
- 150000007949 saponins Chemical class 0.000 description 1
- 201000000306 sarcoidosis Diseases 0.000 description 1
- HFHDHCJBZVLPGP-UHFFFAOYSA-N schardinger α-dextrin Chemical compound O1C(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(O)C2O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC2C(O)C(O)C1OC2CO HFHDHCJBZVLPGP-UHFFFAOYSA-N 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000013207 serial dilution Methods 0.000 description 1
- 239000004208 shellac Substances 0.000 description 1
- 229940113147 shellac Drugs 0.000 description 1
- 235000013874 shellac Nutrition 0.000 description 1
- ZLGIYFNHBLSMPS-ATJNOEHPSA-N shellac Chemical compound OCCCCCC(O)C(O)CCCCCCCC(O)=O.C1C23[C@H](C(O)=O)CCC2[C@](C)(CO)[C@@H]1C(C(O)=O)=C[C@@H]3O ZLGIYFNHBLSMPS-ATJNOEHPSA-N 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 230000005783 single-strand break Effects 0.000 description 1
- 229960005456 sisomicin Drugs 0.000 description 1
- URWAJWIAIPFPJE-YFMIWBNJSA-N sisomycin Chemical compound O1C[C@@](O)(C)[C@H](NC)[C@@H](O)[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H](CC=C(CN)O2)N)[C@@H](N)C[C@H]1N URWAJWIAIPFPJE-YFMIWBNJSA-N 0.000 description 1
- 229960003177 sitafloxacin Drugs 0.000 description 1
- 238000005549 size reduction Methods 0.000 description 1
- 235000019812 sodium carboxymethyl cellulose Nutrition 0.000 description 1
- 229920001027 sodium carboxymethylcellulose Polymers 0.000 description 1
- 235000011069 sorbitan monooleate Nutrition 0.000 description 1
- 239000001593 sorbitan monooleate Substances 0.000 description 1
- 229940035049 sorbitan monooleate Drugs 0.000 description 1
- 235000011076 sorbitan monostearate Nutrition 0.000 description 1
- 239000001587 sorbitan monostearate Substances 0.000 description 1
- 229940035048 sorbitan monostearate Drugs 0.000 description 1
- 235000019337 sorbitan trioleate Nutrition 0.000 description 1
- 229960000391 sorbitan trioleate Drugs 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 229960001294 spiramycin Drugs 0.000 description 1
- 235000019372 spiramycin Nutrition 0.000 description 1
- 229930191512 spiramycin Natural products 0.000 description 1
- 230000003393 splenic effect Effects 0.000 description 1
- 230000002269 spontaneous effect Effects 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 229960004274 stearic acid Drugs 0.000 description 1
- 230000036262 stenosis Effects 0.000 description 1
- 208000037804 stenosis Diseases 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 150000005846 sugar alcohols Polymers 0.000 description 1
- 229960002673 sulfacetamide Drugs 0.000 description 1
- SKIVFJLNDNKQPD-UHFFFAOYSA-N sulfacetamide Chemical compound CC(=O)NS(=O)(=O)C1=CC=C(N)C=C1 SKIVFJLNDNKQPD-UHFFFAOYSA-N 0.000 description 1
- 229960004306 sulfadiazine Drugs 0.000 description 1
- SEEPANYCNGTZFQ-UHFFFAOYSA-N sulfadiazine Chemical compound C1=CC(N)=CC=C1S(=O)(=O)NC1=NC=CC=N1 SEEPANYCNGTZFQ-UHFFFAOYSA-N 0.000 description 1
- 229960000973 sulfadimethoxine Drugs 0.000 description 1
- ZZORFUFYDOWNEF-UHFFFAOYSA-N sulfadimethoxine Chemical compound COC1=NC(OC)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 ZZORFUFYDOWNEF-UHFFFAOYSA-N 0.000 description 1
- 229960002135 sulfadimidine Drugs 0.000 description 1
- 229960000468 sulfalene Drugs 0.000 description 1
- ASWVTGNCAZCNNR-UHFFFAOYSA-N sulfamethazine Chemical compound CC1=CC(C)=NC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 ASWVTGNCAZCNNR-UHFFFAOYSA-N 0.000 description 1
- KXRZBTAEDBELFD-UHFFFAOYSA-N sulfamethopyrazine Chemical compound COC1=NC=CN=C1NS(=O)(=O)C1=CC=C(N)C=C1 KXRZBTAEDBELFD-UHFFFAOYSA-N 0.000 description 1
- 229960005404 sulfamethoxazole Drugs 0.000 description 1
- GPTONYMQFTZPKC-UHFFFAOYSA-N sulfamethoxydiazine Chemical compound N1=CC(OC)=CN=C1NS(=O)(=O)C1=CC=C(N)C=C1 GPTONYMQFTZPKC-UHFFFAOYSA-N 0.000 description 1
- 229960004936 sulfamethoxypyridazine Drugs 0.000 description 1
- VLYWMPOKSSWJAL-UHFFFAOYSA-N sulfamethoxypyridazine Chemical compound N1=NC(OC)=CC=C1NS(=O)(=O)C1=CC=C(N)C=C1 VLYWMPOKSSWJAL-UHFFFAOYSA-N 0.000 description 1
- 229960002229 sulfametoxydiazine Drugs 0.000 description 1
- 229960001363 sulfamoxole Drugs 0.000 description 1
- CYFLXLSBHQBMFT-UHFFFAOYSA-N sulfamoxole Chemical compound O1C(C)=C(C)N=C1NS(=O)(=O)C1=CC=C(N)C=C1 CYFLXLSBHQBMFT-UHFFFAOYSA-N 0.000 description 1
- 229950004215 sulfanitran Drugs 0.000 description 1
- 229960001975 sulfisomidine Drugs 0.000 description 1
- YZMCKZRAOLZXAZ-UHFFFAOYSA-N sulfisomidine Chemical compound CC1=NC(C)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 YZMCKZRAOLZXAZ-UHFFFAOYSA-N 0.000 description 1
- 229940124530 sulfonamide Drugs 0.000 description 1
- 150000003456 sulfonamides Chemical class 0.000 description 1
- JLKIGFTWXXRPMT-UHFFFAOYSA-N sulphamethoxazole Chemical compound O1C(C)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 JLKIGFTWXXRPMT-UHFFFAOYSA-N 0.000 description 1
- 230000000475 sunscreen effect Effects 0.000 description 1
- 239000000516 sunscreening agent Substances 0.000 description 1
- 239000002511 suppository base Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 229960002780 talampicillin Drugs 0.000 description 1
- SOROUYSPFADXSN-SUWVAFIASA-N talampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(=O)OC2C3=CC=CC=C3C(=O)O2)(C)C)=CC=CC=C1 SOROUYSPFADXSN-SUWVAFIASA-N 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
- 239000011975 tartaric acid Substances 0.000 description 1
- 235000002906 tartaric acid Nutrition 0.000 description 1
- XFALPSLJIHVRKE-GFCCVEGCSA-N tedizolid Chemical compound CN1N=NC(C=2N=CC(=CC=2)C=2C(=CC(=CC=2)N2C(O[C@@H](CO)C2)=O)F)=N1 XFALPSLJIHVRKE-GFCCVEGCSA-N 0.000 description 1
- 229960001608 teicoplanin Drugs 0.000 description 1
- LMBFAGIMSUYTBN-MPZNNTNKSA-N teixobactin Chemical compound C([C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(=O)N[C@H](CCC(N)=O)C(=O)N[C@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(=O)N[C@H]1C(N[C@@H](C)C(=O)N[C@@H](C[C@@H]2NC(=N)NC2)C(=O)N[C@H](C(=O)O[C@H]1C)[C@@H](C)CC)=O)NC)C1=CC=CC=C1 LMBFAGIMSUYTBN-MPZNNTNKSA-N 0.000 description 1
- 108010041283 teixobactin Proteins 0.000 description 1
- ONUMZHGUFYIKPM-MXNFEBESSA-N telavancin Chemical compound O1[C@@H](C)[C@@H](O)[C@](NCCNCCCCCCCCCC)(C)C[C@@H]1O[C@H]1[C@H](OC=2C3=CC=4[C@H](C(N[C@H]5C(=O)N[C@H](C(N[C@@H](C6=CC(O)=C(CNCP(O)(O)=O)C(O)=C6C=6C(O)=CC=C5C=6)C(O)=O)=O)[C@H](O)C5=CC=C(C(=C5)Cl)O3)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](NC(=O)[C@@H](CC(C)C)NC)[C@H](O)C3=CC=C(C(=C3)Cl)OC=2C=4)O[C@H](CO)[C@@H](O)[C@@H]1O ONUMZHGUFYIKPM-MXNFEBESSA-N 0.000 description 1
- 229960005240 telavancin Drugs 0.000 description 1
- 108010089019 telavancin Proteins 0.000 description 1
- 229960004576 temafloxacin Drugs 0.000 description 1
- 229960001114 temocillin Drugs 0.000 description 1
- BVCKFLJARNKCSS-DWPRYXJFSA-N temocillin Chemical compound N([C@]1(OC)C(N2[C@H](C(C)(C)S[C@@H]21)C(O)=O)=O)C(=O)C(C(O)=O)C=1C=CSC=1 BVCKFLJARNKCSS-DWPRYXJFSA-N 0.000 description 1
- 150000003505 terpenes Chemical class 0.000 description 1
- IWVCMVBTMGNXQD-UHFFFAOYSA-N terramycin dehydrate Natural products C1=CC=C2C(O)(C)C3C(O)C4C(N(C)C)C(O)=C(C(N)=O)C(=O)C4(O)C(O)=C3C(=O)C2=C1O IWVCMVBTMGNXQD-UHFFFAOYSA-N 0.000 description 1
- 229940040944 tetracyclines Drugs 0.000 description 1
- TUNFSRHWOTWDNC-HKGQFRNVSA-N tetradecanoic acid Chemical compound CCCCCCCCCCCCC[14C](O)=O TUNFSRHWOTWDNC-HKGQFRNVSA-N 0.000 description 1
- OULAJFUGPPVRBK-UHFFFAOYSA-N tetratriacontyl alcohol Natural products CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCO OULAJFUGPPVRBK-UHFFFAOYSA-N 0.000 description 1
- 229940124597 therapeutic agent Drugs 0.000 description 1
- OTVAEFIXJLOWRX-NXEZZACHSA-N thiamphenicol Chemical compound CS(=O)(=O)C1=CC=C([C@@H](O)[C@@H](CO)NC(=O)C(Cl)Cl)C=C1 OTVAEFIXJLOWRX-NXEZZACHSA-N 0.000 description 1
- 229960003053 thiamphenicol Drugs 0.000 description 1
- RYYWUUFWQRZTIU-UHFFFAOYSA-K thiophosphate Chemical compound [O-]P([O-])([O-])=S RYYWUUFWQRZTIU-UHFFFAOYSA-K 0.000 description 1
- 229960004659 ticarcillin Drugs 0.000 description 1
- OHKOGUYZJXTSFX-KZFFXBSXSA-N ticarcillin Chemical compound C=1([C@@H](C(O)=O)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)C=CSC=1 OHKOGUYZJXTSFX-KZFFXBSXSA-N 0.000 description 1
- 229960004089 tigecycline Drugs 0.000 description 1
- 229960005053 tinidazole Drugs 0.000 description 1
- 229960000707 tobramycin Drugs 0.000 description 1
- NLVFBUXFDBBNBW-PBSUHMDJSA-N tobramycin Chemical compound N[C@@H]1C[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N NLVFBUXFDBBNBW-PBSUHMDJSA-N 0.000 description 1
- JOXIMZWYDAKGHI-UHFFFAOYSA-N toluene-4-sulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1 JOXIMZWYDAKGHI-UHFFFAOYSA-N 0.000 description 1
- 229950008187 tosufloxacin Drugs 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 230000007704 transition Effects 0.000 description 1
- 238000013519 translation Methods 0.000 description 1
- 238000011269 treatment regimen Methods 0.000 description 1
- CYRMSUTZVYGINF-UHFFFAOYSA-N trichlorofluoromethane Chemical compound FC(Cl)(Cl)Cl CYRMSUTZVYGINF-UHFFFAOYSA-N 0.000 description 1
- 229940029284 trichlorofluoromethane Drugs 0.000 description 1
- 229940096949 trimethoprim 80 mg Drugs 0.000 description 1
- GPRLSGONYQIRFK-MNYXATJNSA-N triton Chemical compound [3H+] GPRLSGONYQIRFK-MNYXATJNSA-N 0.000 description 1
- 229960005041 troleandomycin Drugs 0.000 description 1
- LQCLVBQBTUVCEQ-QTFUVMRISA-N troleandomycin Chemical compound O1[C@@H](C)[C@H](OC(C)=O)[C@@H](OC)C[C@@H]1O[C@@H]1[C@@H](C)C(=O)O[C@H](C)[C@H](C)[C@H](OC(C)=O)[C@@H](C)C(=O)[C@@]2(OC2)C[C@H](C)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)OC(C)=O)[C@H]1C LQCLVBQBTUVCEQ-QTFUVMRISA-N 0.000 description 1
- 229960000497 trovafloxacin Drugs 0.000 description 1
- WVPSKSLAZQPAKQ-CDMJZVDBSA-N trovafloxacin Chemical compound C([C@H]1[C@@H]([C@H]1C1)N)N1C(C(=CC=1C(=O)C(C(O)=O)=C2)F)=NC=1N2C1=CC=C(F)C=C1F WVPSKSLAZQPAKQ-CDMJZVDBSA-N 0.000 description 1
- 238000004879 turbidimetry Methods 0.000 description 1
- 238000007492 two-way ANOVA Methods 0.000 description 1
- 229960004059 tylosin Drugs 0.000 description 1
- 235000019375 tylosin Nutrition 0.000 description 1
- WBPYTXDJUQJLPQ-VMXQISHHSA-N tylosin Chemical compound O([C@@H]1[C@@H](C)O[C@H]([C@@H]([C@H]1N(C)C)O)O[C@@H]1[C@@H](C)[C@H](O)CC(=O)O[C@@H]([C@H](/C=C(\C)/C=C/C(=O)[C@H](C)C[C@@H]1CC=O)CO[C@H]1[C@@H]([C@H](OC)[C@H](O)[C@@H](C)O1)OC)CC)[C@H]1C[C@@](C)(O)[C@@H](O)[C@H](C)O1 WBPYTXDJUQJLPQ-VMXQISHHSA-N 0.000 description 1
- 241001300301 uncultured bacterium Species 0.000 description 1
- 239000006216 vaginal suppository Substances 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 230000007923 virulence factor Effects 0.000 description 1
- 239000000304 virulence factor Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000012711 vitamin K3 Nutrition 0.000 description 1
- 239000011652 vitamin K3 Substances 0.000 description 1
- 229940041603 vitamin k 3 Drugs 0.000 description 1
- 239000004520 water soluble gel Substances 0.000 description 1
- 229940075420 xanthine Drugs 0.000 description 1
- 239000002676 xenobiotic agent Substances 0.000 description 1
- 229920001221 xylan Polymers 0.000 description 1
- 150000004823 xylans Chemical class 0.000 description 1
- 229950005850 zabofloxacin Drugs 0.000 description 1
- ZNPOCLHDJCAZAH-UCQKPKSFSA-N zabofloxacin Chemical compound CO\N=C1\CN(C=2C(=CC=3C(=O)C(C(O)=O)=CN(C=3N=2)C2CC2)F)CC11CNC1 ZNPOCLHDJCAZAH-UCQKPKSFSA-N 0.000 description 1
- 229940093612 zein Drugs 0.000 description 1
- 239000005019 zein Substances 0.000 description 1
- KGPGQDLTDHGEGT-JCIKCJKQSA-N zeven Chemical compound C=1C([C@@H]2C(=O)N[C@H](C(N[C@H](C3=CC(O)=C4)C(=O)NCCCN(C)C)=O)[C@H](O)C5=CC=C(C(=C5)Cl)OC=5C=C6C=C(C=5O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@H](O5)C(O)=O)NC(=O)CCCCCCCCC(C)C)OC5=CC=C(C=C5)C[C@@H]5C(=O)N[C@H](C(N[C@H]6C(=O)N2)=O)C=2C(Cl)=C(O)C=C(C=2)OC=2C(O)=CC=C(C=2)[C@H](C(N5)=O)NC)=CC=C(O)C=1C3=C4O[C@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]1O KGPGQDLTDHGEGT-JCIKCJKQSA-N 0.000 description 1
- UHVMMEOXYDMDKI-JKYCWFKZSA-L zinc;1-(5-cyanopyridin-2-yl)-3-[(1s,2s)-2-(6-fluoro-2-hydroxy-3-propanoylphenyl)cyclopropyl]urea;diacetate Chemical compound [Zn+2].CC([O-])=O.CC([O-])=O.CCC(=O)C1=CC=C(F)C([C@H]2[C@H](C2)NC(=O)NC=2N=CC(=CC=2)C#N)=C1O UHVMMEOXYDMDKI-JKYCWFKZSA-L 0.000 description 1
- 239000002132 β-lactam antibiotic Substances 0.000 description 1
- 229940124586 β-lactam antibiotics Drugs 0.000 description 1
- 150000003952 β-lactams Chemical class 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/76—Viruses; Subviral particles; Bacteriophages
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
- A61K31/4164—1,3-Diazoles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
- A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/63—Compounds containing para-N-benzenesulfonyl-N-groups, e.g. sulfanilamide, p-nitrobenzenesulfonyl hydrazide
- A61K31/635—Compounds containing para-N-benzenesulfonyl-N-groups, e.g. sulfanilamide, p-nitrobenzenesulfonyl hydrazide having a heterocyclic ring, e.g. sulfadiazine
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
- A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
- A61K31/7036—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin having at least one amino group directly attached to the carbocyclic ring, e.g. streptomycin, gentamycin, amikacin, validamycin, fortimicins
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
- A61K31/7052—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides
- A61K31/7056—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing five-membered rings with nitrogen as a ring hetero atom
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/14—Peptides containing saccharide radicals; Derivatives thereof, e.g. bleomycin, phleomycin, muramylpeptides or vancomycin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/22—Ribonucleases RNAses, DNAses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K48/005—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2310/00—Structure or type of the nucleic acid
- C12N2310/10—Type of nucleic acid
- C12N2310/20—Type of nucleic acid involving clustered regularly interspaced short palindromic repeats [CRISPRs]
Definitions
- the human microbiota comprises bacteria, archaea, viruses, and microbial eukaryotes living in our bodies.
- the taxonomic composition of these communities has been extensively studied and is significantly associated with a variety of diseases and traits.
- This microbiota indeed consists of thousands of different bacterial species that carry hundreds of billions of genes, which is called the microbiome.
- This microbiome encodes for a variety of molecules (proteins, lipids, sugars, RNA, etc.) and functions that are essential and beneficial for their host, for instance the enrichment of glycans metabolism, amino acids and xenobiotics but also the regulation of our immune system. It is also responsible for the synthesis of vitamins, isoprenoids and other nutrients which results in human overall metabolism representing an amalgamation of microbial and human attributes.
- Myocarditis is an inflammatory heart disease that develops into lethal inflammatory cardiomyopathy in 20-30% of the patients (1, 2). Myocarditis can develop into inflammatory cardiomyopathy through chronic stimulation of myosin heavy chain 6-specific Th1 and Th17 cells.
- the mechanisms that govern the cardiotoxicity programming of heart-specific T cells have remained elusive. It is well established that acute immune activation after infectious myocarditis is associated with the generation of autoimmune responses against myosin heavy chain 6 (MYH6) (3-5), while subsequent chronic stimulation of MYH6-specific Th1 and Th17 cells precipitates inflammatory cardiomyopathy (6-9). Nevertheless, it is still unclear which mechanisms mediate the initial activation and cardiotoxicity programming of heart-specific T cells. Likewise, therapeutic approaches that mitigate the activity of such pathogenic T cells and prevent the severe consequences of inflammatory cardiomyopathy are still limited (10, 11).
- a cardinal challenge in deciphering the progressive nature of autoimmune and chronic inflammatory diseases is the deconvolution of their multifactorial nature, which is determined by different degrees of genetic susceptibility and a multitude of environmental conditions (12, 13).
- the quest for genetic determinants underlying susceptibility to myocarditis and dilated cardiomyopathy (DCM) has revealed associations with HLA-DQB1* polymorphisms (14, 15).
- DCM dilated cardiomyopathy
- the invention relates to methods, kits and compositions for reducing the level of or eliminating a Bacteroides bacteria in situ.
- the Bacteroides are killed.
- the invention encompasses methods of preventing myocarditis, treating myocarditis or dilated cardiomyopathy (DCM), or limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in the subject.
- DCM dilated cardiomyopathy
- the invention encompasses methods of diagnosis of a subject as having myocarditis or DCM, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample.
- the invention also encompasses methods of treating myocarditis or DCM in a subject, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample, and when the amount of Bacteroides sp. is higher in the biological sample relative to a control sample, reducing the amount of Bacteroides sp. in the subject.
- the invention encompasses compositions preventing myocarditis, treating myocarditis or DCM, or limiting progression of myocarditis toward DCM in a subject in need thereof.
- the compositions kill Bacteroides bacteria.
- the compositions may reduce the amount of Bacteroides sp. in the subject without killing Bacteroides bacteria.
- the method comprises administering to the subject an effective amount of an antibiotic, bacteria, engineered bacteria, phage, recombinant phage, packaged phagemid, wild-type or synthetic endolysin, wild-type or synthetic bacteriocin or any combination thereof.
- the composition comprises an effective amount of an antibiotic, engineered bacteria, phage, recombinant phage, packaged phagemid, endolysin, bacteriocin or any combination thereof.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the phage, recombinant phage or packaged phagemid encodes an endolysin.
- the phage, recombinant phage, packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- the bacteria or engineered bacteria does not produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which does not produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), still particularly does not produce a ⁇ -galactosidase.
- the Bacteroides is B. thetaiotaomicron and/or B. faecis .
- the Bacteroides is B. thetaiotaomicron and/or B. faecis which produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- a method of diagnosis a subject as having myocarditis or DCM comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample.
- any of these methods comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- the Bacteroides is B. thetaiotaomicron and/or B. faecis.
- a composition for preventing myocarditis in a subject in need thereof comprising a pharmaceutical agent which reduces the amount of Bacteroides sp. in a subject.
- compositions comprising an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- the Bacteroides is B. thetaiotaomicron and/or B. faecis.
- FIGS. 1 A-R Microbiome-dependent transition of autoimmune myocarditis to dilated cardiomyopathy.
- FIG. 1 A Survival of TCRM mice under SPF or GF conditions.
- FIG. 1 B Gross pathology of hearts from 12 week old SPF TCRM and age-matched GF TCRM mice.
- FIG. 1 C Histological analysis of hearts of 12 week old TCRM mice kept under SPF or GF conditions using hematoxylin-eosin (HE) and elastica-van Gieson (EVG) staining.
- FIG. 1 D Histopathological disease severity in TCRM mice under SPF and GF conditions. Dots represent values of individual mice; bar indicates mean disease severity.
- FIG. 1 A Survival of TCRM mice under SPF or GF conditions.
- FIG. 1 B Gross pathology of hearts from 12 week old SPF TCRM and age-matched GF TCRM mice.
- FIG. 1 C Histological analysis of hearts of 12 week old TCRM mice kept under S
- FIG. 1 E-G Echocardiographic parameters in TCRM mice kept under SPF or GF conditions and in transgene-negative littermate of controls with ( FIG. 1 E ) ejection fraction, ( FIG. 1 F ) fractional shortening and ( FIG. 1 G ) systolic left ventricular internal diameter (LVID) determined in individual mice.
- FIG. 1 H-J Co-housing of GF TCRM mice with SPF TCRM mice at the age of 4 and 8 weeks.
- FIG. 1 H Schematic representation of co-housing experiments.
- FIG. 1 I Prospective survival analysis of TCRM mice, arrowheads indicate the age of transfer to SPF conditions.
- FIG. 1 I Prospective survival analysis of TCRM mice, arrowheads indicate the age of transfer to SPF conditions.
- FIG. 1 J Development of myocarditis in TCRM mice under SPF, GF and co-housing conditions; dots represent disease severity in individual mice; bar indicates mean disease severity.
- FIG. 1 K and L Enumeration of heart-infiltrating cells of 12 week old TCRM mice under SPF and GF conditions or GF TCRM mice co-housed (CoH) for 4 weeks in SPF conditions with ( FIG. 1 K ) CD45 + cells and ( FIG. 1 L ) myosin-specific (V ⁇ 8 + CD4 + ) cells.
- FIG. 1 M IFN- ⁇ - and ( FIG. 1 N ) IL-17-producing heart-infiltrating MYH6-specific CD4 + T cells (V ⁇ 8 + CD4 + ).
- FIG. 1 O-R Heart-infiltrating myeloid cell subsets from SPF, GF or 4 week cohoused TCRM mice analyzed by flow cytometry; dots represent values from individual mice, line indicates mean value.
- FIG. 1 A and I ⁇ 12 mice ( FIG. 1 B , C and D), ⁇ 6 mice ( FIG. 1 E-G ), ⁇ 11 mice ( FIG. 1 J ) ⁇ 7 mice ( FIG. 1 M ) per group.
- Statistical analysis was performed using Student's t test ( FIG. 1 D ) or one-way ANOVA with Dunnett's multiple comparison test ( FIG. 1 E-G , J-N and P-R) with *, p ⁇ 0.05; **, p ⁇ 0.01; ***, p ⁇ 0.001.
- FIGS. 2 A-M Interaction of MYH6-specific CD4 + T cells with the intestinal microbiome.
- FIG. 2 A Flow cytometric analysis of mucosal homing markers in heart-infiltrating MYH6 specific cells from SPF or GF TCRM mice.
- FIG. 2 B-C Location and proliferation of CFSE-labelled TCRM cells after adoptive transfer to Rag1 ⁇ / ⁇ mice.
- FIG. 2 B Confocal microscopy analysis of colonic patches at day 3 post adoptive transfer, scale bar 100 ⁇ m ( FIG.
- FIG. 2 C Proliferation of MYH6-specific cells flow cytometry-based quantification of CFSE dilution in the indicated organs and at the indicated time points (mean ⁇ SEM).
- FIG. 2 D Microbiome analysis of 12 week old transgene-negative littermate controls (Tg ⁇ ), SPF TCRM and GF TCRM mice co-housed at 4 weeks of age under SPF conditions.
- FIG. 2 E Principal component analysis of the fecal bacterial composition.
- FIG. 2 F Heat map of the relative abundance of the indicated bacterial classes and families in feces.
- FIG. 2 G-H MYH6-specific CD4 + T cell crossreactivity.
- FIG. 2 G In vitro proliferation of CD4 + T cells from TCRM mice after re-stimulation with MYH6, gal peptide from Bacteroides and cysteine hydrolase-derived peptide from Enterobacter determined by CFSE dilution assay.
- FIG. 2 H Cytokine production of heart- and colon-infiltrating, V ⁇ 8-expressing CD4 + T cells from TCRM mice under SPF conditions after ex vivo re-stimulation with MYH6 peptide and Bacteroides ⁇ -gal peptide (box and whiskers show mean ⁇ interquartile range).
- FIG. 2 I-M GF TCRM mice were monocolonized with parental B.
- FIG. 2 I Schematic representation of the experimental setting. Flow cytometric analysis of ( FIG. 2 J ) heart-infiltrating cells and ( FIG. 2 K ) MYH6-specific cytokine producing cells in the heart and colon (box and whiskers show mean ⁇ interquartile range).
- FIG. 2 L IgA bound fecal bacteria determined by bacterial flow cytometry and ( FIG. 2 M ) pooled data shown as mean ⁇ SEM.
- Statistical analysis was performed using Student's t test ( FIG. 2 A , H, J, K, M) or one-way ANOVA with Dunnett's multiple comparison test ( FIG. 2 F ) with *, p ⁇ 0.05; **, p ⁇ 0.01; ***, p ⁇ 0.001.
- FIGS. 3 A-K Impact of antibiotics treatment on lethal heart disease and immune reactivity in the TCRM model.
- FIG. 3 A Survival
- FIG. 3 B disease severity
- FIG. 3 C histological analysis of hearts of TCRM mice after post-weaning oral treatment with a broad spectrum antibiotics combination comprising of Sulphadoxine, Trimethoprim and Metronidazole (S+T+M); disease severity was determined in 20 week old mice; dots represent values of individual mice; bar indicates mean disease severity.
- FIG. 3 D-J Effect of antibiotics treatment regimen on myocarditis progression in the adoptive transfer myocarditis model.
- FIG. 3 D Schematic representation of the experimental set up.
- FIG. 3 E B. thetaiotaomicron quantification in feces of mice by qPCR in the indicated treatment groups.
- FIG. 3 F Histopathological analysis of hearts from Rag1 ⁇ / ⁇ mice treated with antibiotics at day 28; dots represent values of individual mice; bar indicates mean disease severity. Quantification of heart-infiltrating CD45 + cells ( FIG. 3 G ) and V ⁇ 8-expressing CD4 + T cells ( FIG. 3 H ) in Rag1 ⁇ / ⁇ mice at day 28.
- FIG. 3 I Cytokine production of heart-infiltrating MYH6-specific V ⁇ 8 + CD4 + T cells (box and whiskers show mean ⁇ interquartile range).
- FIG. 3 J Heat map representation of the specific IgG responses against B. thetaiotaomicron, B distasonis, B. vulgatus and E. cloacae determined by ELISA.
- FIG. 3 K Student's t test
- FIG. 3 B Mann-Whitney test
- FIG. 3 E-I one-way ANOVA with Dunnett's multiple comparison test
- FIGS. 4 A-M Immune reactivity against Bacteroides and cardiac myosin antigens in human myocarditis patients.
- FIG. 4 A Analysis of B. thetaiotaomicron -specific IgG antibodies in sera of myocarditis patients from the AMITIS cohort at first admission and different time points post-diagnosis compared to sera from a cohort of healthy individuals, light grey and dark grey squares indicate patients with high or low anti- B. thetaiotaomicron antibody levels respectively, cut value was determined as the mean+2SD of the healthy group; dots represent individual antibody levels, bar indicates mean value.
- FIG. 4 B Ejection fraction (EF) and
- FIG. 4 B Ejection fraction (EF)
- FIG. 4 C C-reactive protein (CRP) values in patients from the AMITIS cohort at the indicated time points; individual values are shown, bar represents mean value.
- FIG. 4 D Composite clinical scores of myocarditis patients from the AMITIS cohort with low vs high IgG antibodies against B. thetaiotaomicron at the first visit (as indicated in A); dots represent individual clinical scores as the sum of positivity for anti-beta1-AR antibodies, CRP values and proportion of EF
- FIG. 4 E-F Anti-bacterial IgG antibody responses in sera of myocarditis patients and healthy controls.
- FIG. 4 E Heat map representation of specific IgG response against B. thetaiotaomicron, B.
- FIG. 4 F B. thetaiotaomicron -specific IgG in serum of patients of the Micro-DCM cohort at admission (individual values are shown, bar indicates mean value) and
- FIG. 4 G heat map representation of antibacterial IgG reactivity in the Micro-DCM cohort.
- FIG. 4 H Heat maps representing the binding of the MYH6 614-629 peptide or the B. thetaiotaomicron ⁇ -gal 11-25 peptide. The in-silico analysis was done for molecules that cover the 99% of the HLA-DQ alleles in open population.
- FIG. 4 J Schematic representation of fecal transplant experiment using stool from two myocarditis patients (HLA-DQB1*03:02 haplotype and B.
- FIG. 4 K B. thetaiotaomicron quantification in feces of TCRM or Tg ⁇ controls at the indicated times post fecal transplantation (box and whiskers show mean ⁇ interquartile range).
- FIG. 4 L-M Enumeration of heart-infiltrating CD45 + immune ( FIG. 4 L ) and CD4 + T cells ( FIG. 4 M ) in recipient mice at 4 weeks after fecal transplantation (mean ⁇ SEM). Statistical analysis was performed using one-way ANOVA with Dunnett's multiple comparison test ( FIG.
- FIGS. 5 A-F Impact of microbial colonization on heart function and inflammation in TCRM mice.
- FIG. 5 A-C Echocardiographic parameters in TCRM mice under SPF, GF and co-housing conditions with ( FIG. 5 A ) ejection fraction, ( FIG. 5 B ) systolic left ventricular internal diameter (LVID) and ( FIG. 5 C ) fractional shortening determined in individual mice.
- FIG. 5 D-F Flow cytometric analysis of heart-infiltrating myeloid cells. Representative dot plots showing CCR2 and Ly6C ( FIG. 5 D ) and CD64 and MHCII expression ( FIG.
- FIG. 5 E shown as percentage of CD11b + Ly6G ⁇ cells depicted for each condition.
- FIGS. 6 A- 1 Activation TCRM CD4 + cells and interaction with the intestinal microbiome.
- FIG. 6 A Schematic representation of the experimental set up for the analysis of homing and early proliferation of CFSE-labelled TCRM cells in Rag ⁇ / ⁇ mice.
- FIG. 6 D Flow cytometric analysis of the proliferation patterns of TCRM cells in the colonic lamina limbal and heart at the indicated time points after a.t.
- FIG. 6 E Cytokine profiles of heart-infiltrating and colonic V ⁇ 8-expressing CD4 + T cells from TCRM mice at the indicated time points analyzed by flow cytometry (mean ⁇ SEM).
- FIG. 6 F Flow cytometry-based cytokine profile analysis of V ⁇ 8-expressing CD4 + T cells from the colonic lamina intestinal of TCRM mice or transgene-negative littermates (Tg ⁇ ).
- FIG. 6 G Production of IFN- ⁇ and IL-17 by colonic CD4 + T cells from TCRM mice or DO11.10 mice after ex-vivo re-stimulation with MYH6 614-629 ; representative dot-plots are shown.
- FIG. 6 H Bacterial ⁇ -diversity in feces of 12 week old TCRM mice and transgene-negative littermates (Tg ⁇ ) under SPF conditions.
- Statistical analysis was performed using Student's t test or one-way ANOVA with Dunnett's multiple comparison test with *, p ⁇ 0.05; **, p ⁇ 0.01; ***, p ⁇ 0.001.
- FIGS. 7 A-I Specificity and immune interaction of Bacteroides mimic peptide-specific CD4 + T cells.
- FIG. 7 A In vitro proliferation of CD4 + T cells from TCRM mice or DO11.10 mice measured by CFSE dilution after re-stimulation with MYH6 or Bacteroides ⁇ -galactosidase peptide or ovalbumin peptide at indicated concentration (representative histograms from 1 out of 2 experiments)
- FIG. 7 B Dendritic cells loaded with MYH6 or ⁇ -gal peptide from Bacteroides at the indicated concentration were co-cultured in vitro with CD4 + T cells from TCRM mice.
- FIG. 7 D - FIG. 7 E Colonization of GF TCRM mice with SPF microbiota or B. thetaiotaomicron or E. coli .
- FIG. 7 F Strategy for the generation of the ⁇ -galactosidase BT1626 deficient B. thetaiotaomicron strain.
- FIG. 7 G Colonization efficiency of WT the parental B. thetaiotaomicron (ATCC29148 ⁇ tdk ) and the ⁇ -gal B.
- FIG. 7 H-I Bacterial flow of fecal IgA bound bacteria in TCRM and Tg ⁇ 12 weeks old mice.
- FIG. 7 H experimental dosing
- Statistical analysis was performed using Student's t test or one-way ANOVA with Dunnett's multiple comparison test with *, p ⁇ 0.05; **, p ⁇ 0.01; ***, p ⁇ 0.001
- FIGS. 8 A-F Effect of antibiotics treatment on splenic and colonic CD4 + T cells.
- FIG. 8 A Microbiota composition analysis by 16S rRNA sequencing of mice treated with a broad spectrum antibiotics cocktail containing sulphadoxine, trimethoprim and metronidazole (S+T+M) compared to untreated TCRM mice.
- FIG. 8 B-F Rag1 ⁇ / ⁇ mice were adoptively transferred with 10 6 splenocytes from TCRM mice and treated with the indicated antibiotics.
- FIG. 8 B Flow cytometry-based enumeration of colonic
- FIG. 8 C V ⁇ 8-expressing CD4 + T and
- FIG. 8 D V ⁇ 8-expressing CD4 + T cells in spleens.
- FIG. 8 E Cytokine production of heart infiltrating V ⁇ 8-expressing CD4 + T cells with representative dot plots.
- FIG. 9 A-C Clinical parameters at admission in B. theta high or low AMITIS patients.
- IgG antibodies against E. cloacae ( FIG. 9 D and FIG. 9 G ), E. coli ( FIG. 9 E and FIG. 9 H ), B. distasonis ( FIG. 9 I ) and B. vulgatus ( FIG. 9 J ) were determined by ELISA and total IgG concentrations ( FIG.
- FIG. 9 F were determined by turbidimetry; dots represent individual patients, bar indicates mean values.
- FIG. 9 K Correlation between MYH6- or ⁇ -gal IFN- ⁇ -producing cells in myocarditis patients of the Micro-DCM cohort and healthy individuals with the indicated HLA-DQB1 alleles; dots represent individual values.
- FIG. 9 L Graphical representation of the multiple risk factors that regulate the outcome of inflammatory cardiomyopathy. Statistical analysis was performed using Student's t test ( FIG. 9 G-J ) or one-way ANOVA with Dunnett's multiple comparison test with ( FIG. 9 D-F ) or Pearson correlation coefficient with two tailed P-value calculation.*, p ⁇ 0.05.
- the present invention unveils the hitherto unknown connection between the microbiome and inflammatory cardiac disease in humans.
- the inventors show here that commensal bacteria such as Bacteroides can serve as a source of a mimic peptide that promotes the progression of myocarditis to inflammatory cardiomyopathy.
- the presented invention also shows that reducing the level of or eliminating Bacteroides in situ with treatment such as antibiotics can prevent lethal heart failure.
- the invention provides the rationale for extended prospective clinical trials to further dissect the connection between the microbiome-driven activation of cross-reactive CD4+ T cells, HLA-dependent genetic predisposition and inflammatory cardiomyopathy.
- Targeting of the microbiome of genetically predisposed myocarditis patients or susceptible patients undergoing checkpoint inhibitor treatment through antibiotics may alleviate disease severity and can therefore contribute to the prevention of the potentially lethal sequelae of inflammatory cardiomyopathy.
- unwanted Bacteroides can be eliminated by using subtractive methods such as antibiotics, phages, recombinant phages, packaged phagemids, wild-type of synthetic endolysins, wild-type of synthetic bacteriocins (such as colicins, pyocins and/or tailocins) which result in the killing of deleterious Bacteroides bacteria or by using competitive methods such as bacteria or engineered bacteria which do not produce said MYH6 mimic peptides and preferably present a competitive advantage over unwanted Bacteroides , which results in the replacement of deleterious Bacteroides bacteria by said non-deleterious bacteria or engineered bacteria.
- Such strategies can significantly reduce the load of Bacteroides populations.
- the invention encompasses compositions, kits and methods for eliminating a Bacteroides bacteria in situ.
- the compositions, kits and methods of the invention eliminate deleterious Bacteroides bacteria, in particular Bacteroides bacteria producing MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly Bacteroides bacteria producing ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), within the host microbiome by killing or reducing the growth of these Bacteroides.
- MYH6 mimic peptides in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17
- Bacteroides bacteria producing ⁇ -gal 11-25 peptide typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22
- the invention encompasses the use of a vector that can transfer with high efficiency a nucleic acid, preferably a plasmid, into a bacterial population within the microbiome that allows the expression of an exogenous enzyme that will modify a gene sequence or directly kill the Bacteroides bacteria.
- the invention further includes methods for screening for elimination of the Bacteroides , for determining the efficiency of vectors at eliminating these Bacteroides , and for determining the effects of these vectors.
- the elimination of Bacteroides bacteria in situ involves the use of antibiotics, of wild-type of synthetic bacteriocins, the use of phages, recombinant phage, packaged phagemid, wild-type of synthetic endolysins, introducing a double strand break in the DNA sequence, or any combination thereof.
- the elimination of Bacteroides bacteria in situ involves the use of bacteria or engineered bacteria, in particular engineered Bacteroides bacteria, which do not produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which do not produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- MYH6 mimic peptides in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17
- ⁇ -gal 11-25 peptide typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22.
- the invention further includes methods for pre-treatment screening of patient to assess the relevancy to treat a patient based on the presence of specific bacteria in the patient intestinal microbiome that produce MYH6 mimic peptides, the genetic background of the patient with the identification of particular set of HLADQB1*03 alleles and inflammatory myocarditis symptoms of the patient.
- the invention further includes a pre-treatment screening of effective antibiotics, effective bacteriocins, effective phages, effective recombinant phages, effective packaged phagemids, effective endolysins, effective bacteria or engineered bacteria or any combination thereof against the targeted Bacteroides bacteria.
- the invention encompasses the use of a vector that can transfer with high efficiency a nucleic acid, preferably a plasmid, into a bacterial population within the microbiome that allows the expression of an exogenous enzyme that will modify a gene sequence or directly kill the Bacteroides bacteria.
- the invention encompasses methods of preventing myocarditis, treating myocarditis or DCM, or limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in the subject.
- the invention encompasses methods of reducing or eliminating a Bacteroides sp. in situ.
- the Bacteroides sp. is reduced or eliminated by contacting it with an antibiotic, for example, as described Gil-Cruz et al., Science 366, 881-886 (2019), which is hereby incorporated by reference.
- the Bacteroides sp. is reduced or eliminated by a genetic modification that leads to the death or reduction in growth of a Bacteroides sp. bacteria, for example, as described in Bikard et al., Cell Host Microbe, Vol. 12, 177-186 (2012) and Bikard et al., Nature Biotechnology Vol. 32 (11) 1146-51 (2014).
- the Bacteroides sp. is reduced or eliminated by contacting it with an endolysin, for example, as described in Gerstmans et al., Biochem Soc Trans. 2016 February; 44(1):123-8, which is hereby incorporated by reference.
- the Bacteroides sp. is reduced or eliminated by contacting it with a bacteriocin (such as a colicin, a pyocin and/or a tailocin).
- a bacteriocin such as a colicin, a pyocin and/or a tailocin.
- the Bacteroides sp. is reduced or eliminated by administering an bacteria or an engineered bacteria, in particular engineered Bacteroides sp. which do not produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which do not produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), and preferably present a competitive advantage over Bacteroides sp. to be reduced or eliminated, for example which comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar.
- engineered bacteria is meant herein a bacteria which has been genetically modified, compared to the wild-type strain, in particular to express proteins of interest and/or to stop expressing unwanted proteins.
- said engineered bacteria comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar
- said engineered bacteria are used in combination with said exogenous food source.
- the method comprises contacting the unwanted bacteria with an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, wild-type or synthetic bacteriocin wild-type or synthetic endolysin or any combination thereof.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the phage, recombinant phage, packaged phagemid encodes a wild-type or synthetic endolysin and/or a wild-type or synthetic bacteriocin
- the phage, recombinant phage, packaged phagemid encodes a nuclease selected from the group consisting of CRISPR-Cas and variants, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- a nuclease selected from the group consisting of CRISPR-Cas and variants, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- the Bacteroides is B. thetaiotaomicron or B. faecis.
- the Bacteroides sp. are contacted in situ with a vector that can transfer with high efficiency a nucleic acid into the bacteria to express an exogenous enzyme (such as Cas9 or Cpf1 also known as Cas12a) in the bacteria that results in bacterial cell death or reduced growth.
- an exogenous enzyme such as Cas9 or Cpf1 also known as Cas12a
- the exogenous enzyme can result in a genetic modification where Cas9 nuclease is used to make the desired modification.
- the invention contemplates introducing a double strand break in the DNA of the bacterial DNA at a specific sequence, for example with a CRISPR/Cas system, together with non-homologous end joining (NHEJ) or homologous recombination (HR) to generate the desired genetic modification.
- NHEJ non-homologous end joining
- HR homologous recombination
- the double strand break is generated in the presence of an editing template comprising homologous region with DNA regions located around the specific sequence located in the bacterial DNA.
- the genetic modification can be a point mutation(s), a deletion(s), insertion(s) or any combination thereof.
- the genetic modification is a point mutation, an insertion or a deletion inside a coding sequence leading to a frameshift mutation or a deletion mutation.
- the genetic modification preferably eliminates, reduces, or increases the expression of a gene.
- the genetic modification can be in the translated or untranslated regions of a gene.
- the genetic modification can be in the promoter region of a gene or within any other region involved in gene regulation.
- the genetic modification integrates a phage genome or exogenous DNA into the host bacterial chromosome or endogenous plasmid(s).
- the genetic modification results in expression of an exogenous protein from an integrated exogenous DNA in the host bacterial chromosome or endogenous plasmid(s). Most preferably, the genetic modification involves either NHEJ or HR endogenous repair mechanism of the host bacteria.
- the genetic modification results in the change in at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 100, 200, 500, etc. amino acids to a different amino acid.
- the genetic modification introduces a stop codon.
- the genetic modification is outside protein coding sequences, within RNA, or within regulatory sequences.
- the genetic modification is within a ⁇ -galactosidase gene, particularly in the coding or non-coding sequence of the ⁇ -galactosidase gene, more particularly in the coding or non-coding sequence of the BT-1626 gene, more particularly in the gene encoding the protein of sequence SEQ ID NO: 1, in particular in the nucleic acid sequence SEQ ID NO: 2.
- Bacteroides sp. can be eliminated by treatment with antibiotic(s).
- the invention encompasses methods of treating a subject with an antibiotic.
- the level of Bacteroides sp. is measured before and after the treatment.
- the invention encompasses a method comprising measuring the level of a Bacteroides sp., subsequently administering an antibiotic, and measuring the level of the Bacteroides sp. after the treatment.
- the antibiotic is streptomycin, vancomycin, clindamycin, or metronidazole, alone or in any possible combination.
- the antibiotic is sulphadoxine, trimethoprim, or metronidazole, alone or in any possible combination, such as described in Gil-Cruz et al., Science 366, 881-886 (2019), which is hereby incorporated by reference.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the antibiotic is selected from the group consisting of penicillins such as penicillin G, penicillin K, penicillin N, penicillin O, penicillin V, methicillin, benzylpenicillin, nafcillin, oxacillin, cloxacillin, dicloxacillin, ampicillin, amoxicillin, pivampicillin, hetacillin, bacampicillin, metampicillin, talampicillin, epicillin, carbenicillin, ticarcillin, temocillin, mezlocillin, and piperacillin; cephalosporins such as cefacetrile, cefadroxil, cephalexin, cephaloglycin, cefalonium, cefaloridine, cefalotin, cefapirin, cefatrizine, cefazaflur, cefazedone, cefazolin, cefradine, cefroxadine, ceftezole, cefaclor, cefonic
- the Bacteroides is resistant to an antibiotic, such as ⁇ -lactams, aminoglycosides, erythromycin or tetracycline.
- an antibiotic such as ⁇ -lactams, aminoglycosides, erythromycin or tetracycline.
- the gene encoding the resistance gene for that antibiotic within the Bacteroides can be modified to make the Bacteroides susceptible to the antibiotic.
- the modified Bacteroides is then treated with the specific antibiotic.
- the elimination of bacteria can be assessed by comparison with and without (control sample) the antibacterial treatment either in vitro or in vivo. Untreated samples can serve as control samples.
- the comparison is preferably performed by assessing the percentage of bacteria before and after the antibacterial treatment at at least two timepoints and determining a reduced amount of the targeted bacteria at later timepoint, for example, as described in the examples.
- Comparison in vitro can be performed by growing the bacteria in solid or liquid culture and determining the percentages or levels of a bacteria over time. The percentages or levels can be determined by routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- Comparison in vivo can be performed by collecting samples (e.g., stool or swab) over time and determining the percentages or levels of a bacteria over time. The percentages can be determined by routine diagnostic procedures employing immunodetection (e.g. ELISA), nucleic acid amplification (e.g., PCR), High Resolution Melting, and nucleic acid sequencing.
- samples e.g., stool or swab
- the percentages can be determined by routine diagnostic procedures employing immunodetection (e.g. ELISA), nucleic acid amplification (e.g., PCR), High Resolution Melting, and nucleic acid sequencing.
- Preferred levels of elimination of bacteria are at least 70%, 80%, 90%, 95%, 97%, 98%, 99%, 99.9%, 99.99%, and 100% of the starting levels of a bacteria.
- the genetic modification is made with one or more of the following enzymes/systems:
- CBE Cytosine base editors
- ABE Adenosine base editors
- Base editors differ in the base modification enzymes.
- CBE rely on ssDNA cytidine deaminase among which: APOBEC1, rAPOBEC1, APOBEC1 mutant or evolved version (evoAPOBEC1), and APOBEC homologs (APOBEC3A (eA3A), Anc689), Cytidine deaminase 1 (CDA1), evoCDA1, FERNY, evoFERNY.
- TadA* is an evolved version of TadA, an E. coli tRNA adenosine deaminase enzyme, able to convert adenosine into Inosine on ssDNA.
- TadA* include TadA-8a-e and TadA-7.10.
- Non-limiting examples of DNA-based editor proteins include BE1, BE2, BE3, BE4, BE4-GAM, HF-BE3, Sniper-BE3, Target-AID, Target-AID-NG, ABE, EE-BE3, YE1-BE3, YE2-BE3, YEE-BE3, BE-PLUS, SaBE3, SaBE4, SaBE4-GAM, Sa(KKH)-BE3, VQR-BE3, VRER-BE3, EQR-BE3, xBE3, Cas12a-BE, Ea3A-BE3, A3A-BE3, TAM, CRISPR-X, ABE7.9, ABE7.10, ABE7.10*, xABE, ABESa, VQR-ABE, VRER-ABE, Sa(KKH)-ABE, ABE8e, SpRY-ABE, SpRY-CBE, SpG-CBE4, SpG-ABE, SpRY-CBE4, SpCas9-NG-ABE, SpCas9-NG-C
- Cytosine Guanine Base Editors consist of a nickase CRISPR fused to:
- CABE Cytosine Adenine Base Editors consist of a Cas9 nickase, a cytidine deaminase (e.g. AID), and a uracil-DNA glycosylase (Ung) (Zhao, D et al. Nature Biotechnology 39:35-40(2021)).
- ACBE include a nucleic acid programmable DNA-binding protein and an adenine oxidase (WO2020181180).
- ATBE consist of a Cas9 nickase and one or more adenosine deaminase or an oxidase domain (WO2020181202).
- TABE consist of a Cas9 nickase and an adenosine methyltransferase, a thymine alkyltransferase, or an adenosine deaminase domain (WO2020181193; WO2020181178; WO2020181195).
- Base editor molecules can also consist of two or more of the above listed editor enzymes fused to a Cas protein (e.g. combination of an ABE and CBE). These biomolecules are named dual base editors and enable the editing of two different bases (Grunewald, J et al. Nature Biotechnology 38:861-864(2020); Li, C et al. Nature Biotechnology 38:875-882(2020)).
- Prime editors consist of a nCas9 fused to a reverse transcriptase used in combination with a prime editing RNA (pegRNA; a guide RNA that includes a template region for reverse transcription).
- pegRNA prime editing RNA
- Prime Editing allows introduction of insertions, deletions (indels), and 12 base-to-base conversions.
- Prime editing relies on the ability of a reverse transcriptase (RT), fused to a Cas nickase variant, to convert RNA sequence brought by a prime editing guide RNA (pegRNA) into DNA at the nick site generated by the Cas protein.
- RT reverse transcriptase
- pegRNA prime editing guide RNA
- the prime editing system can include the expression of an additional sgRNA targeting the Cas nickase activity towards the non-edited DNA strand ideally only after the resolution of the edited strand flap by designing the sgRNA to anneal with the edited strand but not with the original strand.
- Prime editing systems include PE1, PE1-M1, PE1-M2, PE1-M3, PE1-M6, PE1-M15, PE1-M3inv, PE2, PE3, PE3b.
- CRISPEY Cas9 Retron preciSe Parallel Editing via homologY
- SCRIBE strategy a retron system expressed in combination with a recombinase promoting the recombination of single stranded DNA, also known as single stranded annealing proteins (SSAPs) (Farzadfard, F. & Lu, T. K. Science 346, 1256272 (2014)).
- SSAPs single stranded annealing proteins
- Such recombinases include but are not limited to phage recombinases such as lambda red, recET, Sak, Sak4, and newly described SSAPs described in Wannier, T. M. et al. (Improved bacterial recombineering by parallelized protein discovery. Biorxiv 2020.01.14.906594 (2020) doi:10.1101/2020.01.14.906594), which is hereby incorporated by reference.
- the CRISPR system contains two distinct elements, i.e. i) an endonuclease, in this case the CRISPR associated nuclease (Cas or “CRISPR associated protein”) and ii) a guide RNA.
- the guide RNA may be in the form of a chimeric RNA which consists of the combination of a CRISPR (crRNA) bacterial RNA and a tracrRNA (trans-activating RNA CRISPR) 16 .
- the guide RNA combines the targeting specificity of the crRNA corresponding to the “spacing sequences” that serve as guides to the Cas proteins, and the conformational properties of the tracrRNA in a single transcript.
- the target genomic sequence can be permanently interrupted (and causing disappearance of the targeted and surrounding sequences and/or cell death, depending on the location) or modified. The modification may be guided by a repair matrix.
- the CRISPR system includes two main classes depending on the nuclease mechanism of action:
- the sequence of interest according to the present invention comprises a nucleic acid sequence encoding Cas protein.
- CRISPR enzymes are available for use as a sequence of interest on the plasmid according to the present invention.
- the CRISPR enzyme is a Type II CRISPR enzyme, a Type II-A or Type II-B CRISPR enzyme.
- the CRISPR enzyme is a Type I CRISPR enzyme or a Type III CRISPR enzyme.
- the CRISPR enzyme catalyzes DNA modification.
- the CRISPR enzyme catalyzes RNA modification.
- the CRISPR enzymes may be coupled to a guide RNA or single guide RNA (sgRNA).
- the guide RNA or sgRNA targets a gene selected from the group consisting of an antibiotic resistance gene, virulence protein or factor gene, toxin protein or factor gene, a bacterial receptor gene, a membrane protein gene, a structural protein gene, a secreted protein gene, a gene expressing resistance to a drug in general and a gene causing a deleterious effect to the host.
- the CRISPR enzyme makes a double strand break.
- the CRISPR enzyme makes a single strand break or nicks.
- the CRISPR enzyme does not make any break in the DNA or RNA.
- a Cas13-deaminase fusion is used to base edit an RNA.
- the sequence of interest may comprise a nucleic acid sequence encoding a guide RNA or sgRNA to guide the Cas protein endogenous to the targeted bacteria, alone or in combination with a Cas protein and/or a guide RNA encoded by the payload.
- Non-limiting examples of Cas proteins as part of a multi-subunit effector or as a single-unit effector include Cas1, Cas1B, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csn1 and Csx12), Cas10, Cas11 (SS), Cas12a (Cpf1), Cas12b (C2c1), Cas12c (C2c3), Cas12d (CasY), Cas12e (CasX), C2c4, C2c8, C2c5, C2c10, C2c9, Cas13a (C2c2), Cas13b (C2c6), Cas13c (C2c7), Cas13d, Csa5, Csc1, Csc2, Cse1, Cse2, Csy1, Csy2, Csy3, Csf1, Csf2, Csf3, Csf4, Csm1, Csm2, Csm3, C
- the invention encompasses fusion proteins comprising a Cas9 (e.g., a Cas9 nickase) domain and a deaminase domain.
- the fusion protein comprises Cas9 and a cytosine deaminase enzyme, such as APOBEC enzymes, or adenosine deaminase enzymes, such as ADAT enzymes, for example as disclosed in U.S. Patent Publ. 2015/0166980, which is hereby incorporated by reference.
- the deaminase is an ACF1/ASE deaminase.
- the APOBEC deaminase is selected from the group consisting of APOBEC1 deaminase, APOBEC2 deaminase, APOBEC3A deaminase, APOBEC3B deaminase, APOBEC3C deaminase, APOBEC3D deaminase, APOBEC3F deaminase, APOBEC3G deaminase, and APOBEC3H deaminase.
- the fusion protein comprises a Cas9 domain, a cytosine deaminase domain, and a uracil glycosylase inhibitor (UGI) domain.
- the deaminase is an adenosine deaminase that deaminate adenosine in DNA, for example as disclosed in U.S. Pat. No. 10,113,163, which is hereby incorporated by reference.
- the fusion proteins further comprise a nuclear localization sequence (NLS), and/or an inhibitor of base repair, such as, a nuclease dead inosine specific nuclease (dISN), for example as disclosed in U.S. Pat. No. 10,113,163.
- NLS nuclear localization sequence
- DISN nuclease dead inosine specific nuclease
- the invention encompasses fusion proteins comprising a catalytically impaired Cas9 endonuclease fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit, for example as described in Anzalone et al., Nature, Vol. 576, pages 149-157 (2019), which is hereby incorporated by reference.
- pegRNA prime editing guide RNA
- the CRISPR enzyme is any Cas9 protein, for instance any naturally-occurring bacterial Cas9 as well as any variants, homologs or orthologs thereof.
- Cas9 is meant a protein Cas9 (also called Csn1 or Csx12) or a functional protein, peptide or polypeptide fragment thereof, i.e. capable of interacting with the guide RNA(s) and of exerting the enzymatic activity (nuclease) which allows it to perform the double-strand cleavage of the DNA of the target genome.
- “Cas9” can thus denote a modified protein, for example truncated to remove domains of the protein that are not essential for the predefined functions of the protein, in particular the domains that are not necessary for interaction with the gRNA(s).
- the CAS9 is a dCas9 (dead-Cas9) or nCas9 (nickase Cas9) lacking double stranded DNA cleavage activity.
- Cas9 the entire protein or a fragment thereof
- the sequence encoding Cas9 can be obtained from any known Cas9 protein (Fonfara et al., Nucleic Acids Research 42, 2577-2590 (2014); Koonin et al., Current Opinion in Microbiology 37, 67-78 (2017)).
- Cas9 proteins useful in the present invention include, but are not limited to, Cas9 proteins of Streptococcus pyogenes (SpCas9), Streptococcus thermophiles (St1Cas9, St3Cas9), Streptococcus mutans, Staphylococcus aureus (SaCas9), Campylobacter jejuni (CjCas9), Francisella novicida (FnCas9) and Neisseria meningitides (NmCas9).
- SpCas9 Streptococcus pyogenes
- St1Cas9 Streptococcus thermophiles
- St3Cas9 St3Cas9
- Streptococcus mutans Streptococcus mutans
- Staphylococcus aureus SaCas9
- Campylobacter jejuni CjCas9
- Cpf1 (Cas12a) (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cpf1 (Cas12a) protein (Koonin et al., Current Opinion in Microbiology 37, 67-78 (2017)).
- Cpf1 (Cas12a) proteins useful in the present invention include, but are not limited to, Cpf1 (Cas12a) proteins of Acidaminococcus sp, Lachnospiraceae bacteriu and Francisella novicida.
- Cas13a (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cas13a (C2c2) protein (Abudayyeh et al., Nature 550, 280 (2017)).
- Cas13a (C2c2) proteins useful in the present invention include, but are not limited to, Cas13a (C2c2) proteins of Leptotrichia wadei (LwaCas13a).
- Cas13d (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cas13d protein (Yan et al. (2016) Mol Cell 70(2):327-339).
- Cas13d proteins useful in the present invention include, but are not limited to, Cas13d proteins of Eubacterium siraeum and Ruminococcus sp.
- programmable nucleases can be used. These include an engineered TALEN (Transcription Activator-Like Effector Nuclease) and variants, engineered zinc finger nuclease (ZFN) variants, natural, evolved or engineered meganuclease or recombinase variants, and any combination or hybrids of programmable nucleases.
- TALEN Transcription Activator-Like Effector Nuclease
- ZFN zinc finger nuclease
- the programmable nucleases provided herein may be used to selectively modify a Bacteroides sp. DNA encoding a gene of interest.
- RNA base editing is based on the same principle as DNA base editing: an enzyme catalyzing the conversion of a RNA base into another must be brought close to the target base to perform its conversion locally.
- the enzyme used for RNA editing is an adenosine deaminase from ADAR family that converts Adenosine into Inosine in dsRNA structure.
- ADAR DD ADAR deaminase domain
- dPspCas13b catalytically dead Cas13b enzyme fused to a hyperactive mutant of ADAR2 deaminase domain (ADAR2 DD -E488Q for REPAIRv1 and ADAR2 DD -E488Q-T375G for REPAIRv2)
- dPspCas13b catalytically dead Cas13b enzyme fused to a hyperactive mutant of ADAR2 deaminase domain
- Non-limiting examples of RNA based editor proteins include REPAIRv1, REPAIRv2.
- one or more of the following vectors can be used to eliminate or reduce deleterious Bacteroides sp., in particular to introduce the exogenous enzyme that results in a genetic modification or to introduce an endolysin:
- the invention encompasses the use of these vectors wherein the gene editing enzyme/system targets a gene within the target bacteria encoding a protein which is directly or indirectly responsible for a disease or disorder, in particular myocarditis or DCM.
- the gene editing enzyme/system targets a ⁇ -galactosidase gene.
- Bacterial viruses also called bacteriophages or phages
- Bacterial viruses are small viruses displaying the ability to infect and kill bacteria while they do not affect cells from other organisms. Initially described almost a century ago by William Twort, and independently discovered shortly thereafter by Felix d′Herelle, more than 6000 different bacterial viruses have been discovered so far and described morphologically. The vast majority of these viruses are tailed while a small proportion, are polyhedral, filamentous or pleomorphic. They may be classified according to their morphology, their genetic content (DNA vs. RNA), their specific host, the place where they live (marine virus vs. other habitats), and their life cycle.
- phages display different life cycles within the bacterial host: lytic, lysogenic, pseudo-lysogenic, and chronic infection.
- Lytic phages once their DNA injected into their host, replicate their own genome and produce new viral particles at the expense of the host. Indeed, they cause lysis of the host bacterial cell as a normal part of the final stage of their life cycles to liberate viral particles.
- Temperate phages can either replicate by means of the lytic life cycle and cause lysis of the host bacterium, or they can incorporate their DNA into the host bacterial DNA and become non-infectious prophages (lysogenic cycle).
- lytic phages are used.
- a bacteriophage can infect and kill only a small number of different closely-related bacteria.
- packaged phagemids allows to have a defined and controlled way of killing the host.
- Example of packaged phagemids encoding CRISPR-Cas9 or toxins have shown promising results in killing targeted bacterial population (Bikard et al., 2012 , Cell Host & Microbe 12, 177-186; Jiang et al., 2013 , Nat Biotechnol 31, 233-239; Krom et al., 2015 , Nano Letters 15, 4808-4813; Bikard et al, 2014 , Nat Biotech 11, Vol. 32, Citorik, R et al, 2014 , Nat Biotech 11, Vol. 32).
- the vector can comprise a sequence of interest under the control of a promoter.
- the sequence of interest is a programmable nuclease circuit to be delivered to the targeted bacteria.
- This programmable nuclease circuit may be able to mediate in vivo sequence-specific elimination of bacteria that contain a target gene of interest.
- Some embodiments of the present disclosure relate to engineered variants of the Type II CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-associated) system of Streptococcus pyogenes .
- programmable nucleases that can be used include other CRISPR-Cas systems, engineered TALEN (Transcription Activator-Like Effector Nuclease) variants, engineered zinc finger nuclease (ZFN) variants, natural, evolved or engineered meganuclease or recombinase variants, and any combination or hybrids of programmable nucleases.
- CRISPR-Cas systems engineered TALEN (Transcription Activator-Like Effector Nuclease) variants, engineered zinc finger nuclease (ZFN) variants, natural, evolved or engineered meganuclease or recombinase variants, and any combination or hybrids of programmable nucleases.
- TALEN Transcription Activator-Like Effector Nuclease
- ZFN zinc finger nuclease
- sequences of interest preferably programmable, can be added to the payload so as to be delivered to targeted bacteria.
- sequence of interest circuit added to the payload leads to bacteria death.
- the sequence of interest may comprise proteins and enzymes achieving a useful function such as modifying the metabolism of the bacteria, the composition of its environment or affecting the host.
- the nucleic acid sequence of interest is selected from the group consisting of a Cas nuclease, a type I nuclease, a type II nuclease, a type V nuclease, a Cas9 nuclease, a Cpf1 nuclease, a Cas3 nuclease, a Cms1 nuclease, a MAD nuclease, a guide RNA, a single guide RNA (sgRNA), a CRISPR locus, a gene expressing an enzyme such as a nuclease or a kinase, a TALEN, a ZFN, a meganuclease, a recombinase, a bacterial receptor, a membrane protein, a structural protein, a secreted protein, a gene expressing resistance to an antibiotic or to a drug in general, a gene expressing a toxic protein or a toxic factor and
- proteins can also be modified or engineered to include extra features, like the addition or removal of a function (e.g. dCas9), the addition of a secretion signal to a protein not normally secreted, the addition of an exogenous peptide in a loop as non-limiting examples.
- a function e.g. dCas9
- Bacteroides targeted by a composition of the invention can be present in vivo, in a mammalian organism, or in vitro, for example in liquid or solid culture.
- a microbiome may comprise a variety of endogenous Bacteroides species, any of which may be targeted in accordance with the present disclosure.
- the species of targeted Bacteroides bacterial cells may depend on the type of bacteriophages being used for preparing the bacterial virus particles. For example, some bacteriophages exhibit tropism for, or preferentially target, specific host species of bacteria. Other bacteriophages do not exhibit such tropism and may be used to target a number of different genus and/or species of endogenous bacterial cells.
- Bacteroides species include, without limitation, B. acidifaciens , B. barnesiaes, B. caccae , B. caecicola, B. caecigallinarum, B. cellulosilyticus, B. cellulosolvens, B. clarus, B. coagulans, B. coprocola, B. coprophilus, B. coprosuis, B. distasonis, B. eggerthii, B. gracilis, B. faecichinchillae, B. faecis, B. finegoldii, B. fluxus, B. fragilis, B. galacturonicus , B. gallinaceumi, B. gallinarum, B.
- bacterial virus particles may target (e.g., specifically target) a bacterial cell from any one or more of the foregoing species of bacteria to specifically deliver the plasmid according to the invention.
- the targeted bacterial cells are, without limitation, Bacteroides faecis, Bacteroides thetaiotaomicron, Bacteroides fragilis, Bacteroides distasonis, Bacteroides vulgatus , and Bacteroides fragilis.
- the targeted bacterial cells are, without limitation, Bacteroides sp. bacterial which produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which produce ⁇ -galactosidase (typically of sequence SEQ ID NO: 1), more particularly which produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- MYH6 mimic peptides in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17
- ⁇ -galactosidase typically of sequence SEQ ID NO: 1
- ⁇ -gal 11-25 peptide typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22.
- the bacterial virus particles are prepared from bacterial virus.
- the bacterial viruses can be chosen in order to be able to introduce the plasmid into the targeted bacteria.
- Bacterial viruses are preferably bacteriophages. Bacteriophages are obligate intracellular parasites that multiply inside bacteria by co-opting some or all of the host biosynthetic machinery. Phage genomes come in a variety of sizes and shapes (e.g., linear or circular). Most phages range in size from 24-200 nm in diameter. Phages contain nucleic acid (i.e., genome) and proteins, and may be enveloped by a lipid membrane. Depending upon the phage, the nucleic acid genome can be either DNA or RNA, and can exist in either circular or linear forms. The size of the phage genome varies depending upon the phage.
- the simplest phages have genomes that are only a few thousand nucleotides in size, while the more complex phages may contain more than 100,000 nucleotides in their genome, and in rare instances more than 1,000,000.
- the number and amount of individual types of protein in phage particles will vary depending upon the phage.
- Bacteria of the genus Bacteroides can be infected by the following phages: crAss-phage, ad l2, Baf-44, Baf-48B, Baf-64, Bf-I, Bf-52, B40-8, FI, ⁇ I, ⁇ Al, ⁇ BrOI, ⁇ BrO2, 11, 67.1, 67.3, 68.1, mt- Bacteroides (3), Bf42, Bf71, HN-Bdellovibrio (1) and BF-41.
- the vectors disclosed herein may be used in combination with prebiotics.
- Prebiotics include, but are not limited to, amino acids, biotin, fructo-oligosaccharide, galacto-oligosaccharides, hemicelluloses (e.g., arabinoxylan, xylan, xyloglucan, and glucomannan), inulin, chitin, lactulose, mannan oligosaccharides, oligofructose-enriched inulin, gums (e.g., guar gum, gum arabic and carrageenan), oligofructose, oligodextrose, tagatose, resistant maltodextrins (e.g., resistant starch), trans-galactooligosaccharide, pectins (e.g., xylogalacturonan, citrus pectin, apple pectin, and rhamnogalacturonan-I), dietary fiber
- the vectors, antibiotics, bacteriocins and/or endolysins disclosed herein may be used in combination with bacteria and/or engineered bacteria and/or probiotics.
- Probiotics include, but are not limited to lactobacilli, bifidobacteria, streptococci, enterococci, propionibacteria, saccharomycetes, lactobacilli, bifidobacteria, or proteobacteria.
- said bacteria and/or engineered bacteria, in particular engineered Bacteroides sp. and/or probiotics do not produce MYH6 mimic peptides (in particular mimics of MYH6 614-629 or MYH6 614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly do not produce ⁇ -gal 11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), and preferably present a competitive advantage over Bacteroides sp. to be reduced or eliminated.
- said probiotic is an engineered probiotic.
- said engineered bacteria and/or probiotics further comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar.
- said engineered bacteria and/or probiotics may further be engineered to be insensitive to the vectors, phages, antibiotics, bacteriocins and/or endolysins used herein.
- the vector, phage, antibiotic, bacteriocin and/or endolysin disclosed herein is used in combination with (i) an engineered bacteria and/or probiotic further comprising an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source and (ii) said exogenous food source.
- the combined use of the vector disclosed herein and the herein disclosed engineered bacteria and/or probiotic and the corresponding exogenous food source typically enables said engineered bacteria and/or probiotic to fill the niche left empty by the bacteria, in particular the Bacteroides bacteria killed by the use of the vector of the invention.
- Said bacteria and/or engineered bacteria and/or probiotic can typically be administered before, simultaneously with or after said vector, phage, antibiotic, bacteriocin and/or endolysin.
- the invention encompasses methods for screening for genetic modifications in Bacteroides sp.
- the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp., to a subject, subsequently collecting a bacterial sample from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample.
- the genetic modification can be the insertion of an exogenous DNA, for example, encoding an endolysin.
- the method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest.
- the proportion of Bacteroides sp. modified vs non-modified bacteria is quantified.
- Preferred percentages of bacteria with the genetic modification are at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, 99.9%, 99.99%, and 100%.
- the number of non-modified endogenous bacteria is quantified prior to administering a vector.
- the patient can also be pre-screened to determine the genetic signature of the strains the patient carries. This will allow selection of an appropriate capsid to deliver the therapeutic payload based on the genetic signature of the strains the patient carries.
- the vector is in a pharmaceutical or veterinary composition.
- the vector is a bacteriophage.
- the vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- the bacterial sample can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc.
- the level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- the invention encompasses methods for determining the efficiency of a vector at inducing genetic mutations in situ.
- the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a naturally occurring bacteria, to a subject, subsequently collecting a bacterial sample from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest and quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample.
- the vector is in a pharmaceutical or veterinary composition.
- the vector is a bacteriophage.
- the vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- the bacterial sample can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc.
- the level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- the invention encompasses methods for determining the effect of a genetic mutation on bacterial growth.
- the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp., to a subject, subsequently collecting at least two sequential bacterial samples from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest and quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest in said bacterial samples.
- the vector is in a pharmaceutical or veterinary composition.
- the vector is a bacteriophage.
- the vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- the bacterial samples can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc.
- the samples can be collected at any sequential time points. Preferably, the time between these collections is at least 3, 6, 12, 24, 48, 72, 96 hours or 7, 14, 30, 60, 120, or 365 days.
- the level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- All of the screening methods of the invention can use any of the vectors and enzymes/systems of the invention to screen for any of the genetic modification of the invention.
- All of the screening methods of the invention can further include a step of comparing the level of bacteria containing a genetic modification in a base of a DNA of interest with the level of bacteria not containing a genetic modification the base of a DNA of interest in a bacterial sample.
- All of the screening methods of the invention can further include a step of contacting the vector with bacteria in liquid or solid culture and quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample.
- the method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest.
- the method comprises providing a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp.
- the method can further comprise contacting the vector with a Bacteroides sp. in liquid or solid culture and quantitating the level of Bacteroides sp. containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample.
- the method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest.
- the levels of bacteria containing a genetic modification in a base of a DNA of interest can be compared with the level of bacteria not containing a genetic modification the base of a DNA of interest over time in the culture. Preferably, the time between these comparisons is at least 1, 2, 3, 4, 5, 6, 12, 24, 48, 72, or 96 hours.
- the invention encompasses methods for determining the efficiency of a bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin at reducing or killing Bacteroides sp. in situ.
- the method comprises providing a bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin, contacting the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin with a Bacteroides sp. in situ, and quantitating the level of Bacteroides sp. killed or reduced by the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin.
- the endolysin or bacteriocin is encoded by a vector, such as a bacteriophage.
- the method can further comprise quantitating the level of Bacteroides not killed or reduced by the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin.
- the levels of bacteria can be compared over time. Preferably, the time between these comparisons is at least 1, 2, 3, 4, 5, 6, 12, 24, 48, 72, or 96 hours.
- the invention encompasses pharmaceutical and veterinary compositions comprising the vectors, bacteria, engineered bacteria, antibiotics, bacteriocins and/or endolysins of the invention.
- the invention encompasses a pharmaceutical agent which reduces the amount of Bacteroides sp. in a subject.
- the invention encompasses in situ administration of the pharmaceutical or veterinary composition to the bacteria in a subject. Any method known to the skilled artisan can be used to contact the composition with the bacterial target in situ.
- the composition comprises an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
- the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- the phage, recombinant phage, packaged phagemid encodes an endolysin or a bacteriocin.
- the phage, recombinant phage, packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- the Bacteroides is B. thetaiotaomicron or B. faecis.
- the pharmaceutical or veterinary composition according to the invention may further comprise a pharmaceutically acceptable vehicle.
- a solid pharmaceutically acceptable vehicle may include one or more substances which may also act as flavoring agents, lubricants, solubilizers, suspending agents, dyes, fillers, glidants, compression aids, inert binders, sweeteners, preservatives, dyes, coatings, or tablet-disintegrating agents.
- Suitable solid vehicles include, for example calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidone, low melting waxes and ion exchange resins.
- the pharmaceutical or veterinary composition may be prepared as a sterile solid composition that may be suspended at the time of administration using sterile water, saline, or other appropriate sterile injectable medium.
- the pharmaceutical or veterinary compositions of the invention may be administered orally in the form of a sterile solution or suspension containing other solutes or suspending agents (for example, enough saline or glucose to make the solution isotonic), bile salts, acacia, gelatin, sorbitan monooleate, polysorbate 80 (oleate esters of sorbitol and its anhydrides copolymerized with ethylene oxide) and the like.
- the particles according to the invention can also be administered orally either in liquid or solid composition form.
- compositions suitable for oral administration include solid forms, such as pills, capsules, granules, tablets, and powders, and liquid forms, such as solutions, syrups, elixirs, and suspensions.
- forms useful for enteral administration include sterile solutions, emulsions, and suspensions.
- the bacterial virus particles according to the invention may be dissolved or suspended in a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fats.
- a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fats.
- the liquid vehicle can contain other suitable pharmaceutical additives such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavouring agents, suspending agents, thickening agents, colours, viscosity regulators, stabilizers or osmo-regulators.
- suitable examples of liquid vehicles for oral and enteral administration include water (partially containing additives as above, e.g. cellulose derivatives, preferably sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols, e.g.
- the vehicle can also be an oily ester such as ethyl oleate and isopropyl myristate.
- Sterile liquid vehicles are useful in sterile liquid form compositions for enteral administration.
- the liquid vehicle for pressurized compositions can be a halogenated hydrocarbon or other pharmaceutically acceptable propellant.
- the invention encompasses pharmaceutical or veterinary composition formulated for delayed or gradual enteric release.
- formulations or pharmaceutical preparations of the invention are formulated for delivery of the vector into the distal small bowel and/or the colon.
- the formulation can allow the vector to pass through stomach acid and pancreatic enzymes and bile, and reach undamaged to be viable in the distal small bowel and colon.
- the pharmaceutical or veterinary composition is micro-encapsulated, formed into tablets and/or placed into capsules, preferably enteric-coated capsules.
- the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release, using cellulose acetate (CA) and polyethylene glycol (PEG).
- the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using a hydroxypropylmethylcellulose (HPMC), a microcrystalline cellulose (MCC) and magnesium stearate.
- the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using e.g., a poly(meth)acrylate, e.g. a methacrylic acid copolymer B, a methyl methacrylate and/or a methacrylic acid ester, or a polyvinylpyrrolidone (PVP).
- the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using a release-retarding matrix material such as: an acrylic polymer, a cellulose, a wax, a fatty acid, shellac, zein, hydrogenated vegetable oil, hydrogenated castor oil, polyvinylpyrrolidone, a vinyl acetate copolymer, a vinyl alcohol copolymer, polyethylene oxide, an acrylic acid and methacrylic acid copolymer, a methyl methacrylate copolymer, an ethoxyethyl methacrylate polymer, a cyanoethyl methacrylate polymer, an aminoalkyl methacrylate copolymer, a poly(acrylic acid), a poly(methacrylic acid), a methacrylic acid alkylamide copolymer, a poly(methyl methacrylate), a poly(methacrylic acid anhydride), a methyl methacrylate polymer, a polymethacrylate polymer
- the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release as described in U.S. Pat. App. Pub. 20110218216, which describes an extended release pharmaceutical composition for oral administration, and uses a hydrophilic polymer, a hydrophobic material and a hydrophobic polymer or a mixture thereof, with a microenvironment pH modifier.
- the hydrophobic polymer can be ethylcellulose, cellulose acetate, cellulose propionate, cellulose butyrate, methacrylic acid-acrylic acid copolymers or a mixture thereof.
- the hydrophilic polymer can be polyvinylpyrrolidone, hydroxypropylcellulose, methylcellulose, hydroxypropylmethyl cellulose, polyethylene oxide, acrylic acid copolymers or a mixture thereof.
- the hydrophobic material can be a hydrogenated vegetable oil, hydrogenated castor oil, carnauba wax, candelilla wax, beeswax, paraffin wax, stearic acid, glyceryl behenate, cetyl alcohol, cetostearyl alcohol or and a mixture thereof.
- the microenvironment pH modifier can be an inorganic acid, an amino acid, an organic acid or a mixture thereof.
- the microenvironment pH modifier can be lauric acid, myristic acid, acetic acid, benzoic acid, palmitic acid, stearic acid, oxalic acid, malonic acid, succinic acid, adipic acid, sebacic acid, fumaric acid, maleic acid; glycolic acid, lactic acid, malic acid, tartaric acid, citric acid, sodium dihydrogen citrate, gluconic acid, a salicylic acid, tosylic acid, mesylic acid or malic acid or a mixture thereof.
- the pharmaceutical or veterinary compositions are a powder that can be included into a tablet or a suppository.
- a formulation or pharmaceutical preparation of the invention can be a ‘powder for reconstitution’ as a liquid to be drunk or otherwise administered.
- the pharmaceutical or veterinary compositions can be administered in a cream, gel, lotion, liquid, feed, or aerosol spray.
- a bacteriophage is immobilized to a solid surface using any substance known in the art and any technology known in the art, for example, but not limited to immobilization of bacteriophages onto polymeric beads using technology as outlined in U.S. Pat. No. 7,482,115, which is incorporated herein by reference. Phages may be immobilized onto appropriately sized polymeric beads so that the coated beads may be added to aerosols, creams, gels or liquids.
- the size of the polymeric beads may be from about 0.1 ⁇ m to 500 ⁇ m, for example 50 ⁇ m to 100 ⁇ m.
- the coated polymeric beads may be incorporated into animal feed, including pelleted feed and feed in any other format, incorporated into any other edible device used to present phage to the animals, added to water offered to animals in a bowl, presented to animals through water feeding systems.
- the compositions are used for treatment of surface wounds and other surface infections using creams, gels, aerosol sprays and the like.
- the pharmaceutical or veterinary compositions can be administered by inhalation, in the form of a suppository or pessary, topically (e.g., in the form of a lotion, solution, cream, ointment or dusting powder), epi- or transdermally (e.g., by use of a skin patch), orally (e.g., as a tablet, which may contain excipients such as starch or lactose), as a capsule, ovule, elixirs, solutions, or suspensions (each optionally containing flavoring, coloring agents and/or excipients), or they can be injected parenterally (e.g., intravenously, intramuscularly or subcutaneously).
- parenterally e.g., intravenously, intramuscularly or subcutaneously.
- compositions may be used in the form of a sterile aqueous solution which may contain other substances, for example enough salts or monosaccharides to make the solution isotonic with blood.
- compositions may be administered in the form of tablets or lozenges which can be formulated in a conventional manner.
- a bacteriophage and/or polypeptide of the present invention is administered topically, either as a single agent, or in combination with other antibiotic treatments, as described herein or known in the art.
- the pharmaceutical or veterinary compositions can also be dermally or transdermally administered.
- the pharmaceutical or veterinary composition can be combined with one or a combination of carriers, which can include but are not limited to, an aqueous liquid, an alcohol base liquid, a water soluble gel, a lotion, an ointment, a nonaqueous liquid base, a mineral oil base, a blend of mineral oil and petrolatum, lanolin, liposomes, proteins carriers such as serum albumin or gelatin, powdered cellulose carmel, and combinations thereof.
- a topical mode of delivery may include a smear, a spray, a bandage, a time-release patch, a liquid-absorbed wipe, and combinations thereof.
- the pharmaceutical or veterinary composition can be applied to a patch, wipe, bandage, etc., either directly or in a carrier(s).
- the patches, wipes, bandages, etc. may be damp or dry, wherein the phage and/or polypeptide (e.g., a lysin) is in a lyophilized form on the patch.
- the carriers of topical compositions may comprise semi-solid and gel-like vehicles that include a polymer thickener, water, preservatives, active surfactants, or emulsifiers, antioxidants, sun screens, and a solvent or mixed solvent system.
- U.S. Pat. No. 5,863,560 discloses a number of different carrier combinations that can aid in the exposure of skin to a medicament, and its contents are incorporated herein.
- the pharmaceutical or veterinary composition is conveniently delivered in the form of a dry powder inhaler or an aerosol spray presentation from a pressurized container, pump, spray, or nebuliser with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, a hydrofluoroalkane such as 1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane, carbon dioxide, or other suitable gas.
- a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, a hydrofluoroalkane such as 1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane, carbon dioxide, or other suitable gas.
- the dosage unit may be determined by
- the pressurized container, pump, spray, or nebuliser may contain a solution or suspension of the active compound, e.g., using a mixture of ethanol and the propellant as the solvent, which may additionally contain a lubricant, e.g., sorbitan trioleate.
- a lubricant e.g., sorbitan trioleate.
- Capsules and cartridges made, for example, from gelatin) for use in an inhaler or insufflator may be formulated to contain a powder mix of the bacteriophage and/or polypeptide of the invention and a suitable powder base such as lactose or starch.
- compositions of the invention can be applied topically in the form of a gel, hydrogel, lotion, solution, cream, ointment, or dusting powder.
- Compositions of the invention may also be administered by the ocular route.
- the compositions of the invention can be formulated as micronized suspensions in isotonic, pH adjusted, sterile saline, or, preferably, as solutions in isotonic, pH adjusted, sterile saline, optionally in combination with a preservative such as a benzalkonium chloride.
- they may be formulated in an ointment such as petrolatum.
- Dosages and desired drug concentrations of the pharmaceutical and veterinary composition compositions of the present invention may vary depending on the particular use. The determination of the appropriate dosage or route of administration is within the skill of an ordinary physician. Animal experiments can provide reliable guidance for the determination of effective doses in human therapy.
- the pharmaceutical or veterinary composition can be formulated into ointment, cream or gel form and appropriate penetrants or detergents could be used to facilitate permeation, such as dimethyl sulfoxide, dimethyl acetamide and dimethylformamide.
- nasal sprays for transmucosal administration, nasal sprays, rectal or vaginal suppositories can be used.
- the active compounds can be incorporated into any of the known suppository bases by methods known in the art. Examples of such bases include cocoa butter, polyethylene glycols (carbowaxes), polyethylene sorbitan monostearate, and mixtures of these with other compatible materials to modify the melting point or dissolution rate.
- the subject according to the invention is an animal, preferably a mammal, even more preferably a human.
- the term “subject” can also refer to non-human animals, in particular mammals such as dogs, cats, horses, cows, pigs, sheep, donkeys, rabbits, ferrets, gerbils, hamsters, chinchillas, rats, mice, guinea pigs and non-human primates, among others, or non-mammals such as poultry, that are in need of treatment.
- the human subject according to the invention may be a human at the prenatal stage, a new-born, a child, an infant, an adolescent or an adult at any age.
- the subject is carrying a HLA-DQ haplotype which binds to human MYH6 614629 (typically of sequence SEQ ID NO: 17); more particularly (i) a HLA-DQB1*03 variant (in particular encoded by DQB1*03:01, DQB1*03:02, DQB1*03:03, DQB1*03:04, or DQB1*03:05 alleles) and/or (ii) a HLA-DQA1*01:02 or a HLA-DQA1*05:01 variant; still particularly a HLA-DQ7 (for example encoded by DQA1*02:01/DQB1*03:01, DQA1*03:01/DQB1*03:01, DQA1*03:03/DQB1*03:01, DQA1*03:01/DQB1*03:04, DQA1*03:02/DQB1*03:04, DQA1*03:02/
- the subject has been diagnosed with, or is at risk of developing myocarditis or DCM. Diagnostic methods of determining Bacteroides sp. infection are well known by the man skilled in the art.
- the subject has never received any treatment prior to the administration of the vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention.
- the subject has already received at least one line of treatment, preferably several lines of treatment, prior to the administration of the vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention.
- the subject suffers from cancer.
- the subject has already received an immune-checkpoint inhibitor treatment, more particularly a combination of anti-CTLA-4 and anti-PD-1 therapy, in particular to treat cancer.
- the subject suffers from an auto-immune disease, in particular from sarcoidosis, rheumatoid arthritis, dermatomyositis, systemic lupus erythematosus (SLE) or giant cell myocarditis.
- an auto-immune disease in particular from sarcoidosis, rheumatoid arthritis, dermatomyositis, systemic lupus erythematosus (SLE) or giant cell myocarditis.
- the subject suffers from systemic lupus erythematosus (SLE). More particularly, the subject carries a mutation in the gene encoding the 3′-5′ DNA exonuclease TREX1.
- the subject suffers from a bacterial, viral, parasital, protozoan or fungal infection, which may cause myocarditis.
- the subject has already received an antiretroviral treatment, an antipsychotic treatment, an anticancer treatment and/or anti-parasite treatment, such as an anti-malaria treatment.
- the treatment is administered regularly, preferably between every day and every month, more preferably between every day and every two weeks, more preferably between every day and every week, even more preferably the treatment is administered every day.
- the treatment is administered several times a day, preferably 2 or 3 times a day, even more preferably 3 times a day.
- the duration of treatment with vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention is preferably comprised between 1 day and 20 weeks, more preferably between 1 day and 10 weeks, still more preferably between 1 day and 4 weeks, even more preferably between 1 day and 2 weeks.
- the duration of the treatment is of about 1 week.
- the treatment may last as long as the infection, disorder and/or disease persists.
- the form of the pharmaceutical or veterinary compositions, the route of administration and the dose of administration of vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably of a payload according to the invention, particularly of a payload packaged into a delivery vehicle according to the invention, preferably of a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention can be adjusted by the man skilled in the art according to the type and severity of the infection (e.g. depending on the bacteria species involved in the disease, disorder and/or infection and its localization in the patient's or subject's body), and to the patient or subject, in particular its age, weight, sex, and general physical condition.
- the amount of vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention, to be administered has to be determined by standard procedure well known by those of ordinary skills in the art. Physiological data of the patient or subject (e.g. age, size, and weight) and the routes of administration have to be taken into account to determine the appropriate dosage, so as a therapeutically effective amount will be administered to the patient or subject.
- the total amount of delivery vehicles particularly a payload packaged into a delivery vehicle according to the invention, preferably a plasmid or phagemid packaged into a bacterial virus particle according to the invention, for each administration is comprised between 10 4 and 10 15 delivery vehicles.
- vector refers to any construct of sequences that are capable of expression of a polypeptide in a given host cell. If a vector is used then the choice of vector is dependent upon the method that will be used to transform host bacteria as is well known to those skilled in the art.
- Vectors can include, without limitation, plasmid vectors and recombinant phage vectors, or any other vector known in that art suitable for delivering a polypeptide of the invention to target bacteria.
- the skilled artisan is well aware of the genetic elements that must be present on the vector in order to successfully transform, select and propagate host cells comprising any of the isolated nucleotides or nucleic acid sequences of the invention.
- delivery vehicle refers to any vehicle that allows the transfer of a payload into a bacterium.
- delivery vehicle encompassed by the present invention including, without limitation, bacteriophage scaffold, virus scaffold, bacterial virus particle, chemical based delivery vehicle (e.g., cyclodextrin, calcium phosphate, cationic polymers, cationic liposomes), protein-based or peptide-based delivery vehicle, lipid-based delivery vehicle, nanoparticle-based delivery vehicles, non-chemical-based delivery vehicles (e.g., transformation, electroporation, sonoporation, optical transfection), particle-based delivery vehicles (e.g., gene gun, magnetofection, impalefection, particle bombardment, cell-penetrating peptides) or donor bacteria (conjugation).
- chemical based delivery vehicle e.g., cyclodextrin, calcium phosphate, cationic polymers, cationic liposomes
- protein-based or peptide-based delivery vehicle e.g., lipid-based delivery vehicle, nanoparticle-based delivery vehicles, non-chemical-based delivery vehicles (e.
- the delivery vehicle can refer to a bacteriophage derived scaffold and can be obtained from a natural, evolved or engineered capsid.
- the delivery vehicle is the payload as bacteria are naturally competent to take up a payload from the environment on their own.
- the delivery vehicle is a bacterial virus particle, in particular a packaged phagemid.
- the delivery vehicle in particular the bacteriophage, bacterial virus particle or packaged phagemid, comprising the vector of the invention is incapable of self-reproduction.
- self-reproduction is different from “self-replication”, “self-replication” referring to the capability of replicating a nucleic acid, whereas “self-reproduction” refers to the capability of having a progeny, in particular of producing new delivery vehicles, said delivery vehicles being either produced empty or with a nucleic acid of interest packaged.
- delivery vehicle incapable of self-reproduction is meant herein that at least one, several or all functional gene(s) necessary to produce said delivery vehicle is(are) absent from said delivery vehicle (and from said vector included in said delivery vehicle).
- said at least one, several or all functional gene(s) necessary to produce said delivery vehicle is(are) present in the donor cell as defined above, preferably in a plasmid or in a helper phage present in the donor cell as defined above, enabling the production of said delivery vehicle in said donor cell.
- said functional gene necessary to produce delivery vehicle may be absent through (i) the absence of the corresponding gene or (ii) the presence of the corresponding gene but in a non-functional form.
- sequence of said gene necessary to produce said delivery vehicle is absent from said delivery vehicle.
- sequence of said gene necessary to produce said delivery vehicle has been replaced by a nucleic acid sequence of interest, in particular by a nucleic acid sequence encoding enzymes or systems for inducing genetic modifications, as defined above.
- said gene necessary to produce said delivery vehicle is present in said delivery vehicle in a non-functional form, for example in a mutant non-functional form, or in a non-expressible form, for example with deleted or mutated non-functional regulators.
- said gene necessary to produce said delivery vehicle is present in said delivery vehicle in a mutated form which renders it non-functional in the target cell, while remaining functional in the donor cell.
- genes necessary to produce said delivery vehicle encompass any coding or non-coding nucleic acid required for the production of said delivery vehicle.
- genes necessary to produce said delivery vehicle include genes encoding phage structural proteins; phage genes involved in the control of genetic expression; phage genes involved in transcription and/or translation regulation; phage genes involved in phage DNA replication; phage genes involved in production of phage proteins; phage genes involved in phage proteins folding; phage genes involved in phage DNA packaging; and phage genes encoding proteins involved in bacterial cell lysis.
- the term «payload» refers to any nucleic acid sequence or amino acid sequence, or a combination of both (such as, without limitation, peptide nucleic acid or peptide-oligonucleotide conjugate) transferred into a bacterium with a delivery vehicle.
- payload may also refer to a plasmid, a vector or a cargo.
- the payload can be a phagemid or phasmid obtained from natural, evolved or engineered bacteriophage genome.
- the payload can also be composed only in part of phagemid or phasmid obtained from natural, evolved or engineered bacteriophage genome.
- the payload is the delivery vehicle as bacteria are naturally competent to take up a payload from the environment on their own.
- nucleic acid refers to a sequence of at least two nucleotides covalently linked together which can be single-stranded or double-stranded or contains portion of both single-stranded and double-stranded sequence.
- Nucleic acids of the present invention can be naturally occurring, recombinant or synthetic.
- the nucleic acid can be in the form of a circular sequence or a linear sequence or a combination of both forms.
- the nucleic acid can be DNA, both genomic or cDNA, or RNA or a combination of both.
- the nucleic acid may contain any combination of deoxyribonucleotides and ribonucleotides, and any combination of bases, including uracil, adenine, thymine, cytosine, guanine, inosine, xanthine, hypoxanthine, isocytosine, 5-hydroxymethylcytosine and isoguanine.
- bases including uracil, adenine, thymine, cytosine, guanine, inosine, xanthine, hypoxanthine, isocytosine, 5-hydroxymethylcytosine and isoguanine.
- modified bases that can be used in the present invention are detailed in Chemical Reviews 2016, 116 (20) 12655-12687.
- nucleic acid also encompasses any nucleic acid analogs which may contain other backbones comprising, without limitation, phosphoramide, phosphorothioate, phosphorodithioate, O-methylphosphoroamidite linkage and/or deoxyribonucleotides and ribonucleotides nucleic acids. Any combination of the above features of a nucleic acid is also encompassed by the present invention.
- phagemid or “phasmid” are equivalent and refer to a recombinant DNA vector comprising at least one sequence of a bacteriophage genome and which is preferably not able of producing progeny, more particularly a vector that derives from both a plasmid and a bacteriophage genome.
- a phagemid of the disclosure comprises a phage packaging site and optionally an origin of replication (ori), in particular a bacterial and/or phage origin of replication.
- the phagemid does not comprise a bacterial origin of replication and thus cannot replicate by itself once injected into a bacterium.
- the phagemid comprises a plasmid origin of replication, in particular a bacterial and/or phage origin of replication.
- the term “packaged phagemid” refers to a phagemid which is encapsidated in a bacteriophage scaffold, bacterial virus particle or capsid. Particularly, it refers to a bacteriophage scaffold, bacterial virus particle or capsid devoid of a bacteriophage genome.
- the packaged phagemid may be produced with a helper phage strategy, well known from the man skilled in the art.
- the helper phage comprises all the genes coding for the structural and functional proteins that are indispensable for the phagemid according to the invention to be encapsidated.
- the packaged phagemid may be produced with a satellite virus strategy, also known from the man skilled in the art.
- Satellite virus are subviral agent and are composed of nucleic acid that depends on the co-infection of a host cell with a helper virus for all the morphogenetic functions, whereas for all its episomal functions (integration and immunity, multicopy plasmid replication) the satellite is completely autonomous from the helper.
- the satellite genes can encode proteins that promote capsid size reduction of the helper phage, as described for the P4 Sid protein that controls the P2 capsid size to fit its smaller genome.
- peptide refers both to a short chain of at least 2 amino acids linked between each other and to a part of, a subset of, or a fragment of a protein which part, subset or fragment being not expressed independently from the rest of the protein.
- a peptide is a protein.
- a peptide is not a protein and peptide only refers to a part, a subset or a fragment of a protein.
- the peptide is from 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 amino acids to 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 40, 50, 100, 200 amino acids in size.
- TCR-M MYH6-specific TCR transgenic mice (TCR-M) on the BALB/c background have been previously described (8).
- TCRM mice were maintained as heterozygous and transgene-negative littermates were used as controls.
- TCRM mice were re-derived to germ-free conditions by axenic 2-cell stage embryo transfer into pseudopregnant germ-free recipient females at the Clean Mouse Facility, University of Berne, Switzerland, and bred and maintained under germ-free conditions in flexible film isolators. Germ-free status was routinely monitored by culture-dependent (aerobic and anerobic) and -independent (Sytox DNA stain of cecal contents) methods and were additionally confirmed pathogen-free.
- mice germ-free TCRM mice were transferred at 4 or 8 weeks of age to the SPF facility in the Institute of Immunobiology, Kantonsspital St. Gallen and co-housed with SPF TCRM mice and transgene-negative littermates.
- Rag1 ⁇ / ⁇ and DO11.10 mice were obtained from the Jackson laboratories.
- Homozygous OVA-TCR transgenic mice (DO11.10) were mated with SPF BALB/c Thy1.1 mice from the TCRM colony of the St. Gallen facility and offspring were used for analysis.
- Rag1 ⁇ / ⁇ mice were co-housed with TCRM mice 3 weeks before set-up of the breeding pairs. All mice were on the BALB/c genetic background and were maintained in individually ventilated cages.
- Myocarditis patients with typical late enhancement pattern in cardiac MRI and exclusion of coronary artery disease by angiography or computed tomography.
- Heart failure patients with left ventricular ejection fraction ⁇ 40%, no obvious mechanism underlying left ventricular dysfunction and no significant coronary artery by invasive angiography (no stenosis >50% in any main vessel).
- Histological analysis was performed as previously described (8). Briefly, hearts were fixed in 4% formaldehyde (Formafix) for at least 12 h and embedded in paraffin. Histopathological changes were evaluated following hematoxylin/eosin (HE) and Elastica van Gieson (EVG) staining. Myocarditis severity was evaluated using a semiquantitative scoring system: 0, no inflammation; 1, ⁇ 100 inflammatory cells involved, small inflammatory lesions; 2, >100 inflammatory cells involved, larger inflammatory lesions; 3, >10% of the heart section involved in inflammation; 4, >30% of the heart section involved in inflammation; 5, >30% of the heart section involved in inflammation with extensive fibrosis and dilation of ventricle. Images from heart sections were acquired using a Leica DMRA microscope.
- mice received 2 ⁇ g of anti-CD45.2 BV510 or anti-Thy 1.1 PE i.v. 5 min before being euthanized in the indicated experiments.
- Euthanized animals were perfused with 20 ml PBS and small heart tissue pieces were placed into a 6-well dish filled with RPMI 1640 medium containing 2% FCS, 20 mM HEPES (Lonza), 1 mg/ml collagenase D (Sigma) and 25 ⁇ g/ml DNasel (Applichem) and incubated at 37° C. under continuous stirring.
- tissue pieces were mechanically disrupted and mononuclear cells were purified by centrifugation (25 min at 800 ⁇ g, 4° C.) on a 30-70% Percoll gradient (GE Healthcare).
- GE Healthcare Percoll gradient
- colons were flushed and tissue was harvested and incubated three times for 15 min at room temperature under constant agitation with BSS containing 5% FCS (Lonza), 5 mM EDTA (Sigma), 10 mM HEPES (Sigma) and 1 mM DTT in order to dissociate the epithelial layer.
- BSS containing 10 mM HEPES and digested three times for 20 min at 37° C.
- Murine colonic tissue and lymph nodes were fixed overnight at 4° C. in freshly prepared 4% paraformaldehyde (Merck Millipore) under agitation. Tissues were embedded and oriented in 4% low melting agarose (Invitrogen) in PBS and serially sectioned with a vibratome (Leica VT-1200). 30-40 ⁇ m thick sections were blocked in PBS containing 10% FCS, 1 mg/ml anti-Fc ⁇ receptor (BD Biosciences) and 0.1% Triton X-100 (Sigma). Tissues were incubated overnight at 4° C. with the following antibodies: anti-mouse CD4, anti-mouse E-Cadherin (Biolegend). Microscopy was performed using a confocal microscope (LSM-710, Carl Zeiss). Microscopy data were recorded with ZEN 2010 software (Carl Zeiss, Inc.) and processed in Imaris Versions 7 (Bitplane).
- lymphocytes were incubated for 3.5 h at 37° C. in 96-well round-bottom plates in 200 ⁇ L of RPMI per 5% FCS supplemented with 10 ⁇ g/mL brefeldin A (Sigma).
- Cells were stimulated with 0.25 ⁇ g of the different peptides (see below), or phorbol myristate acetate (50 ng/ml; Sigma)/ionomycin (500 ng/ml; Sigma) (PMA/I) as positive control, or were left untreated. After surface molecule labelling, cells were stained with the fixable viability dye Zombie Aqua (Biolegend).
- cytofix-cytoperm (BD Biosciences) for 20 min. Fixed cells were incubated at 4° C. for 40 min with permeabilization buffer (2% FCS, 0.5% saponin in PBS) containing antibodies to CD3, CD4, IFN- ⁇ and IL-17A. Samples were measured using a BD LSR Fortessa (BD Biosciences). Data were analyzed using FlowJo software (Tree Star, Inc.).
- Bt_bgal_ko_1 AAG ATA ACA TTC GAG TOG Forward primer 1 kb upstream ACT ATT TCG GCA ATA ATA of BT1626, with Gibson CGC TGA AAG (SEQ ID NO: overhang homologous to 3) pExchange-tdk.
- Bt_bgal_ko_2 CAA ATG CAA TCA GTT TCA Reverse primer immediately GGC ATT ATT ATA TTG TTT upstream of BT1626, with TTT GGT GAC TGG T (SEQ Gibson overhang homologous ID NO: 4) to the downstream region of BT1626.
- the plasmid was then transformed into the conjugative E. coli strain SM10 and conjugated into the B. thetaiotaomicron ⁇ tdk.
- the resulting colonies are resistant to erythromycin and have the complete plasmid inserted into the chromosome resulting from a single homologous recombination event.
- Counterselection on plates containing 200 ⁇ g/ml 5FUdR led to a second recombination event, and either to a reversion to the wild type state or to the deletion of BT1626.
- Successful deletion was tested using PCR with control primers, and clones with a clean deletion of the gene were further used ( FIG. 7 F and Table 6).
- All plates used for generating mutants were Brain Heart Infusion (Oxoid), supplemented with 2% agar (Sigma), 10% defibrinated sheep blood (Sigma), 1 ⁇ g/ml Hemin (Sigma). 200 ⁇ g/ml FUdR (Sigma), 25 ⁇ g/ml erythromycin (Sigma), and 200 ⁇ g/ml gentamicin (Sigma) were added before pouring the plates when needed.
- the different Bacteroides thetaiotaomicron strains (ATCC29148, ATCC29148 ⁇ tdk or ATCC29148 ⁇ tdk ⁇ BT1626) were grown anaerobically overnight at 37° C. in Brain Heart Infusion (BHI, CM1135, Oxoid) with the addition of hemin (1 ⁇ g/ml, Sigma) and menadione (0.5 ⁇ g/ml, Sigma).
- E. coli BL21(DE3) was grown in LB medium at 37° C. Overnight broth culture of B. thetaiotaomicron or E.
- coli was washed once in PBS and gavaged intra-gastrically (500 ⁇ l/mouse corresponding to 10 9 bacteria/mouse) to germ-free mice (at least 3-4 week old).
- Germ-free and monocolonized animals were kept in flexible film isolators until the end of the experiments and monitored regularly for excluding any contaminations using Sytox staining on fecal pellets for GF animals, Gram staining and 16S rRNA sequencing on fecal pellets for monocolonized animals (34).
- the effects of mono-colonization in TCRM mice were analyzed 8 weeks post colonization.
- Spleens were collected from TCRM mice and disrupted on a 70- ⁇ m cell strainer. Red blood cells were lysed by osmotic shock and 10 6 splenocytes were injected intravenously in 4 wk mice in the lateral tail vein of Rag1 ⁇ / ⁇ mice. Seven days after adoptive transfer, mice were bled to confirm CD4 + T cell expansion. Disease activity scores and analysis of T cell activation in the heart and colonic laminalitis were performed at day 28 post adoptive transfer.
- mice were treated for 4 or 16 weeks with the following antibiotics in the drinking water (i) a broad spectrum antibiotic mixture; Sulfadoxine 400 mg/l and Trimethoprim 80 mg/l (Borgal; Veterinaria) or (ii) Streptomycin 200 mg/l (Sigma-Aldrich) to reduce the proportion of Bacteroidetes (35) or iii) Vancomycin 100 mg/l.
- Metronidazole Sigma-Aldrich
- Spleen cells were labelled with 10 ⁇ l of 5 mM carboxyfluorescein succinimidyl ester (CSFE, dissolved in DMSO) (Molecular Probes) in 10 ml of PBS for 10 min at 37° C. The staining reaction was stopped with 1 mL FCS (Lonza), followed by washing with PBS.
- CSFE carboxyfluorescein succinimidyl ester
- a total of 2 ⁇ 10 5 splenocytes/well were seeded in 96-well round-bottom plate and MYH6 614-629 (RSLKLMATLFSTYASADR, SEQ ID NO: 10) or Bacteroides ⁇ -galactosidase 11-25 (TFLILMAALTATFASAQK, SEQ ID NO: 11), Enterobacter cysteine hydrolase 10-27 (LLIGMMSTFSTYASAQET, SEQ ID NO: 12) or OVA 323-339 (ISQAVHAAHAEINEAGR, SEQ ID NO: 13) peptide was added at the indicated concentrations.
- CSFE dilution was assessed by flow cytometry after 3 days of incubation at 37° C. All peptides were synthetized by Genscript with >80% of purity.
- Spleen cells were labelled with CFSE as described above and 5 ⁇ 10 6 cells were i.v. transferred in to Rag ⁇ / ⁇ mice. Mice were culled at days 2, 3, 4, or 7 after adoptive transfer and the presence of myosin specific (CD4 + V ⁇ 2 + V ⁇ 8 + ) cells as well as CFSE dilution was assessed by flow cytometry in single cell suspensions from the mediastinal lymph node, colonic lymph node, colonic lamina intestinal and heart.
- High-binding 96-well polystyrene plates (Corning) were coated with 10 7 CFU of the following bacterial strains B. theta ATCC29148, B. distasionis, B. vulgatus, E. cloacae ATCC 13047 or E. coli K12 in 0.1 M carbonate-bicarbonate buffer, pH 9.5. Plates were incubated for 1 h at 37° C. and then overnight at 4° C. Before use, plates were washed three times with PBS containing 0.05% Tween-20 (PBS-T) (Sigma-Aldrich). Non-specific binding was blocked with 5% non-fat dry milk diluted in PBS (PBS-M) for 1 h at 37° C.
- PBS-T PBS containing 0.05% Tween-20
- Optical density was measured at 492 nm using an automated ELISA plate reader (Tecan). Absorbance values at 1:160 dilutions are presented in the figures.
- high and low responders to B. thetaiotaomicron were high responders were defined as patients that had an IgG at diagnosis of A >mean of healthy controls +2 SD.
- Fecal-bound IgA was assessed using bacterial flow cytometry for IgA-bound bacteria performed as previously described (36). In brief, fecal pellets were homogenized in 1 ml of staining buffer (PBS+1% (w/v) bovine serum albumin (BSA, Sigma). Tubes were centrifuged at 400 g for 5 minutes to pellet large debris and the supernatant was filtered through a sterile 70 ⁇ m cell strainer and transferred to a new tube.
- staining buffer PBS+1% (w/v) bovine serum albumin
- Samples were centrifuged at 8000 ⁇ g for 5 minutes to pellet bacteria and pellets were re-suspended in PBS with SYTO BC (1:1000, Life Technologies) for 15 minutes on ice, Samples were washed with PBS and stained in 100 ⁇ l staining buffer containing PE-conjugated anti-mouse IgA (1:75) for 30 minutes on ice. Samples were washed 3 times with PBS and re-suspended for flow cytometry analysis in a FACS CANTO (BD). Bacteria were gated based on forward and side scatter and the gating strategy was verified by SYTO-BC staining. Fecal samples from Rag1 ⁇ / ⁇ mice were used as negative controls to define the threshold for IgA ⁇ and IgA + populations. Analysis of IgA binding was done using FlowJo (v10.1).
- TCRM SPF mice For microbial composition analysis, the feces from TCRM SPF mice, Tg ⁇ littermates and TCRM GF co-housed with TCRM SPF and Tg ⁇ littermates were used. Microbial composition was assessed by a 16S high-throughput amplicon analysis as described previously (37).
- the 163 rRNA gene segments spanning the variable V5 and V6 regions were amplified from DNA from the samples, using a multiplex approach with the barcoded forward fusion primer 5′-CCATCTCATCCCTGCGTGTCTCCGACTCAG (SEQ ID NO:14) BARCODE ATTAGATACCCYGGTAGTCC-3′ (SEQ ID NO: 23) in combination with the reverse fusion primer 5′-CCTCTCTATGGGCAGTCGGTGATACGAGCTGACGACARCCATG-3′ (SEQ ID NO: 15).
- the PCR-amplified 16S V5-V6 amplicons were purified and prepared for sequencing on the Ion torrent PGM system according to the manufacturers instructions (Life Technologies). Samples with over 10′000 reads were accepted for analysis.
- Bacterial genomic DNA was extracted using a commercial kit (QIAamp DNA Stool Mini Kit) with minor modifications. Glass beads (Sigma, G4649-10G) were used for an optimal disruption of fecal content and an additional extraction step was introduced after incubation of bacteria with a lysis buffer (20 mg/ml lysozyme—Sigma 62970-5G-F, 2 mM EDTA, 1.2% Triton, 20 mM Tris-HCI). Extracted genomic DNA was used as a template for B. thetaiotaomicron PCR amplification and quantification using a real-time PCR machine (Quant Studio 5) and SYBR Green PCR technology (Applied Biosystems, Thermo Fisher Scientific). B.
- thetaiotaomicron was quantified using species specific primers that have been established and validated before (39) (atcgcaaaaataagatgggcaaa and cacaacagccatagcgttcca) with a cycling protocol of denaturation at 95° C. (2 min), 40 cycles of 95° C. (20 sec), 58° C. (20 sec) and 72° C. (30 sec), Each 17 ⁇ l qPCR mixture consisted of 8.5 ⁇ l of 2 ⁇ SYBR Green MasterMix (Applied Biosystems, Thermo Fisher Scientific), 1.7 ⁇ l of BSA (100 ⁇ g/ml), 0.6 ⁇ l of each primer (10 ⁇ M), 3.60 ⁇ l PCR-grade water and 2 ⁇ l of extracted genomic DNA.
- species specific primers that have been established and validated before (39) (atcgcaaaaataagatgggcaaa and cacaacagccatagcgttcca) with a cycling protocol of denaturation at 95° C. (2
- DNA extracted from B. theta ATCC29148 was used to construct a standard curve with 10-fold dilutions of DNA templates of known concentrations. Concentrations of DNA used in the standard curves ranged from 33.75 ng/ ⁇ l equivalent to 10 7 bacteria to 0.035 pg/ ⁇ l equivalent to 10 1 bacteria. The PCR amplifications were performed in duplicates and the detection limit was 0.34 pg/ ⁇ l (10 2 bacteria). Bacterial qPCR signals were normalized to 1 ⁇ g of total bacterial DNA.
- Proteins encoded in cultured and uncultured bacteria of the NCBI microbial protein database were searched for 15 amino acid sequences similar to MYH6 614-629 with BLAST p2.3.1 (blast.ncbi.nlm.nih.gov).
- Predicted binding of the candidate bacterial peptides to the BALB/c mouse IA d MHC class II was evaluated using the immune epitope database and analysis resource IEDB from the National Institute of Allergy and Infectious diseases online prediction tool (tools.iedb.org/mhcii/).
- the binding threshold was set at 10, and peptides with percentile ranks lower than the threshold value were predicted to be good MHC II haplotype binders (Table 1).
- the HLA-genotyping for the AMITIS patients was performed by the Institut für Klinische Transfusionstechnik and Hämotherapie from the (2015)sklinikum Wurzburg. Briefly, genomic DNA was obtained from PBMCs and analyzed by a sequence based typing (SBT) by Sanger sequencing using forward and reverse primers for sequencing the exons 2 and 3 (HLA class H) in a Applied Biosystems Genetic analyzer 3130 ⁇ I, instrument (ThermoFisher Scientific), using SBTexcellerator kits (Gendx) and the analysis software SBTengine.
- SBT sequence based typing
- Gendx SBTexcellerator kits
- the Micro-DCM cohort HLA typing was performed by Illumina sequencing (Histogenetics USA).
- Dynabeads® M-270 Epoxy (Life Technologies) coated for 72 h according to the manufacturer's instructions with 100, 80, 60, 40, 20, 10, 5 or 2.5 ⁇ g/ml ⁇ 1-ECII peptide.
- Control beads were coated with 10 ⁇ g/ml of a scramble peptide comprising the same amino acids of the ⁇ 1-ECII peptide. After the coating the beads were washed using phosphate-buffered saline (PBS)/0.1% Bovine Serum Albumin (BSA) and stored in PBS/0.1% BSA/0.02% NaN3 until further use.
- PBS phosphate-buffered saline
- BSA Bovine Serum Albumin
- the beads were washed four times using PBS/0.1% BSA/0.02% NaN3 and transferred to fresh wells before incubation with a 1:100 dilution of the secondary antibody F(ab)2 donkey-anti-human IgG (H+L) RTC (Dianova) in PBS/0.1% BSA/0.02% NaN3 (total volume: 50 ⁇ l, 15 min, on ice). 13F6 was detected using F(ab)2 donkey-anti-rat IgG (H+L) PE (Dianova; 1:1000 dilution in PBS/0.1% BSA/0.02% NaN3). After a final washing step the samples were measured on a FACScan flow cytometer and the data analyzed using FlowJo (Treestar). Half maximal binding was determined using GraphPad Prism. When a half maximal binding value was calculable, the serum was defined as antibody-positive.
- IFN- ⁇ ELISPOT was performed as described by Lv et. al. with minor modifications (7). Briefly, PBMCs from patients and healthy volunteers were isolated on density gradients. PBMCs were adjusted to 2 ⁇ 10 6 cells/ml in 0.5 ml of complete IVS medium supplemented with 8% of human serum. Cells were stimulated for 48 h with 0.25 ⁇ g/ml of the indicated peptide or left untreated as medium control. After incubation, cells were washed and re-suspended in 0.3 ml of RPMI/10% FCS and approximately 5 ⁇ 10 5 cells/0.1 ml were dispensed in duplicates into previously coated and blocked ELISPOT plates (IFN- ⁇ base kit, MABTECH).
- ELISPOT plates were washed and developed as indicated by the manufacturer. Plates were counted using an ELISPOT reader and analyzed with the software ELISPOT 3.1SR (AID). Number of specific IFN- ⁇ positive cells was calculated as the mean of the duplicates minus the mean of the medium control.
- Fecal microbiota transplantation was performed as previously reported by Gopalakrishnan et. al (41). Briefly, germ free TCRM and transgene-negative littermates received FMT from myocarditis patients which were previously confirmed to be positive for B. thetaiotaomicron and the DQB1*03:02 HLA allele; each patient sample was administered to 4 mice. 200 ⁇ l cleared supernatant from 0.1 g/ ⁇ l human fecal suspension was obtained using a 100 ⁇ m strainer and gavaged into mice 3 times over 1 week. B. thetaiotaomicron colonization was assessed by qPCR at 14 and 21 days after the first administration.
- FIG. 1 D Echocardiographic analysis of heart functions with recording of the ejection fraction ( FIG. 1 E ), fractional shortening ( FIG. 1 F ) and systolic left ventricle internal diameter (LVID) ( FIG. 1 G ) revealed that hearts of 12 week old GF TCRM mice functioned normally despite the low level cardiac inflammation.
- TCRM mice were transferred from GF to the SPF environment at 4 and 8 weeks of age and co-housed (CoH) with SPF TCRM mice ( FIG. 1 H ). Colonization with the SPF microbiome precipitated progression to lethal disease in ex-GF mice within 6-12 weeks ( FIG. 1 I ), significantly exacerbated cardiac inflammation ( FIG. 1 J ) and impaired cardiac function ( FIG. 5 A-C ). Moreover, microbial colonization of GF TCRM mice led to rapid infiltration of the heart with CD45 + immune cells ( FIG.
- FIG. 1 K IFN- ⁇ production of heart-infiltrating TCRM Th cells did not differ between SPF and GF mice
- FIG. 1 N the expression of IL-17 was dependent on the microbial status
- Co-housing increased the fraction of both IFN- ⁇ -( FIG. 1 M ) and IL-17-secreting heart-specific CD4 + T cells ( FIG. 1 N ) and promoted the accumulation of inflammatory myeloid cells in the heart tissue ( FIG. 1 O-R and FIG. 5 D-F ) including inflammatory monocytes expressing CCR2 ( FIG.
- IL-17 determines the severity of experimental myocarditis in mice (6, 8) and appears to promote heart failure in human myocarditis patients (9). Since balancing of Th17 cell activity is crucial for the integrity and immune homeostasis of the intestinal lamina propria (26), we hypothesized that MYH6-specific CD4 + T cells in TCRM mice receive remote activation signals from the intestinal microbiota. Indeed, assessment of mucosal homing molecule expression on heart-infiltrating CD4 + TCRM cells revealed significantly elevated integrin ⁇ 4 ⁇ 7 and chemokine receptor CCR6 expression under SPF compared to GF conditions ( FIG. 2 A ).
- FIG. 6 A Adoptive transfer of dye-labeled TCRM cells into Rag1-deficient BALB/c mice revealed homing of CD4 + cells to the T cell zones of colonic patches ( FIG. 2 B ), colonic lymph nodes ( FIG. 6 B ) and mesenteric lymph nodes ( FIG. 6 C ).
- FIG. 2 D Next generation sequencing of 16S rRNA genes amplified from feces revealed that co-housed Tg ⁇ and TCRM mice possess disparate microbiomes ( FIG. 6 H ) and that the microbiome of co-housed ex-GF TCRM mice appears to be more similar to the SPF TCRM microbiome ( FIG. 2 E ). More detailed analyses confirmed that GF TCRM mice acquired the microbiome of SPF TCRM mice ( FIG.
- TCRM CD4 + T cells exhibit a functional avidity of 6.3 ⁇ 10 ⁇ 6 M for the cross-reactive B.
- theta peptide FIG. 7 B
- B theta peptide that facilitated efficient activation of TCRM T cells in vivo by B.
- theta peptide-pulsed dendritic cells FIG. 6 C
- ex vivo re-stimulation of transgenic CD4 + T cells from heart and colon FIG.
- FIG. 7 F Mono-colonization of GF TCRM mice with B. theta ⁇ -gal or the parental strain ( FIG. 2 I ) resulted in efficient seeding of the intestinal microenvironments with both strains ( FIG. 7 G ), but precipitated significantly reduced accumulation of CD45 + immune cells and V ⁇ 8 + CD4 + T cells in the myocardium ( FIG. 2 J ). Moreover, IL-17 production of myosin-specific CD4 + T cells in heart and colon was significantly reduced when TCRM mice were re-colonized with B. theta ⁇ -gal ( FIG.
- FIG. 3 G with significantly reduced accumulation of TCR-transgenic CD4 + T cells in hearts of Rag1 ⁇ / ⁇ recipients treated with Streptomycin, S+T+M or Vancomycin ( FIG. 3 H ).
- these three antibiotic regimens reduced CD45 + immune cell ( FIG. 8 B ) and V ⁇ 8 + CD4 + T cell accumulation in the colon ( FIG. 8 C ), but not in the spleen of Rag1 ⁇ / ⁇ recipients ( FIG. 8 D ).
- Vancomycin and S+T+M antibiotics treatment had a pronounced impact on the activation of heart-infiltrating and colonic Th17 cells, while the activity of Th1 cells was reduced to a lesser extent ( FIG. 3 I and FIG. 8 E-F ).
- the adoptively transferred B cells from TCRM spleens generated B. theta -specific IgG antibodies that could be detected in the serum of Rag1 ⁇ / ⁇ recipients on day 28 post transfer ( FIG. 3 J ).
- Antibiotics treatment reduced B. theta -specific IgG antibodies, but had variable effects on IgG antibody responses against B. distasonis, B. vulgatus or Enterobacter cloacae ( FIG. 3 J ) indicating that antibiotics treatment did not negatively impact global immune reactivity in the Rag1 ⁇ / ⁇ recipients.
- MYH6 is the dominant autoantigen in the heart tissue (5, 28) and that susceptibility to experimental autoimmune myocarditis depends mainly on the MHC haplotype, whereby MYH6 epitopes appear to be presented by different MHC class II molecules (29).
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Molecular Biology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- Immunology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Gastroenterology & Hepatology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Mycology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Virology (AREA)
- Cardiology (AREA)
- Heart & Thoracic Surgery (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention relates to methods, kits and compositions for reducing the level of or eliminating Bacteroides in situ. The invention encompasses methods of preventing myocarditis, treating myocarditis or dilated cardiomyopathy, or limiting progression of myocarditis toward dilated cardiomyopathy in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in the subject. The invention further encompasses methods of diagnosis of a subject as having myocarditis or dilated cardiomyopathy. The invention also encompasses compositions preventing myocarditis, treating myocarditis or dilated cardiomyopathy, or limiting progression of myocarditis toward dilated cardiomyopathy in a subject in need thereof.
Description
- This application is a divisional of U.S. application Ser. No. 17/225,854, filed Apr. 8, 2021, which claims the benefit of U.S. application 63/007,197 filed Apr. 8, 2020, which are herein incorporated by reference in its entirety.
- The contents of the electronic sequence listing (EB2020-03USDiv_SequenceListing.xml; Size: 26,978 bytes; and Date of Creation: Jan. 24, 2023) is herein incorporated by reference in its entirety.
- The human microbiota comprises bacteria, archaea, viruses, and microbial eukaryotes living in our bodies. The taxonomic composition of these communities has been extensively studied and is significantly associated with a variety of diseases and traits. This microbiota indeed consists of thousands of different bacterial species that carry hundreds of billions of genes, which is called the microbiome. This microbiome encodes for a variety of molecules (proteins, lipids, sugars, RNA, etc.) and functions that are essential and beneficial for their host, for instance the enrichment of glycans metabolism, amino acids and xenobiotics but also the regulation of our immune system. It is also responsible for the synthesis of vitamins, isoprenoids and other nutrients which results in human overall metabolism representing an amalgamation of microbial and human attributes.
- Interestingly, there is a growing body of recent studies based on metagenome sequencing that demonstrate that the presence of specific strains (and therefore of specific genetic signatures in the microbiome) can be directly linked to a number of pathologies, including auto-immunity, infections, inflammation or tumorigenesis.
- Myocarditis is an inflammatory heart disease that develops into lethal inflammatory cardiomyopathy in 20-30% of the patients (1, 2). Myocarditis can develop into inflammatory cardiomyopathy through chronic stimulation of myosin heavy chain 6-specific Th1 and Th17 cells. However, the mechanisms that govern the cardiotoxicity programming of heart-specific T cells have remained elusive. It is well established that acute immune activation after infectious myocarditis is associated with the generation of autoimmune responses against myosin heavy chain 6 (MYH6) (3-5), while subsequent chronic stimulation of MYH6-specific Th1 and Th17 cells precipitates inflammatory cardiomyopathy (6-9). Nevertheless, it is still unclear which mechanisms mediate the initial activation and cardiotoxicity programming of heart-specific T cells. Likewise, therapeutic approaches that mitigate the activity of such pathogenic T cells and prevent the severe consequences of inflammatory cardiomyopathy are still limited (10, 11).
- A cardinal challenge in deciphering the progressive nature of autoimmune and chronic inflammatory diseases is the deconvolution of their multifactorial nature, which is determined by different degrees of genetic susceptibility and a multitude of environmental conditions (12, 13). The quest for genetic determinants underlying susceptibility to myocarditis and dilated cardiomyopathy (DCM) has revealed associations with HLA-DQB1* polymorphisms (14, 15). Moreover, the development of progressive myocarditis in transgenic mice expressing the HLA-DQ8 haplotype (encoded by the DQA1*03:01/DQB1*03:02 alleles) (7, 16) indicates that antigen presentation via certain MHC class II molecules is a major determinant of myocarditis.
- A variety of pathogens including enteroviruses, Borrelia burgdorferi and Trypanosoma cruzi can cause cardiac damage leading to myocarditis thereby predisposing affected patients for inflammatory cardiomyopathy (11). Still, the question whether and to which extent pathogen-induced death of cardiomyocytes results in the excessive presentation of self-antigens to MHC class II-restricted T cells or whether antigenic mimicry of microbial components drives the disease has remained unanswered (10, 11). The present invention fulfills this need.
- The invention relates to methods, kits and compositions for reducing the level of or eliminating a Bacteroides bacteria in situ. Preferably, the Bacteroides are killed.
- The invention encompasses methods of preventing myocarditis, treating myocarditis or dilated cardiomyopathy (DCM), or limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in the subject.
- The invention encompasses methods of diagnosis of a subject as having myocarditis or DCM, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample.
- The invention also encompasses methods of treating myocarditis or DCM in a subject, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample, and when the amount of Bacteroides sp. is higher in the biological sample relative to a control sample, reducing the amount of Bacteroides sp. in the subject.
- The invention encompasses compositions preventing myocarditis, treating myocarditis or DCM, or limiting progression of myocarditis toward DCM in a subject in need thereof. Preferably, the compositions kill Bacteroides bacteria. Alternatively, the compositions may reduce the amount of Bacteroides sp. in the subject without killing Bacteroides bacteria.
- In various embodiments, the method comprises administering to the subject an effective amount of an antibiotic, bacteria, engineered bacteria, phage, recombinant phage, packaged phagemid, wild-type or synthetic endolysin, wild-type or synthetic bacteriocin or any combination thereof.
- In various embodiments, the composition comprises an effective amount of an antibiotic, engineered bacteria, phage, recombinant phage, packaged phagemid, endolysin, bacteriocin or any combination thereof.
- In various embodiments, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- In various embodiments, the phage, recombinant phage or packaged phagemid encodes an endolysin.
- In various embodiments, the phage, recombinant phage, packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- In various embodiments, the bacteria or engineered bacteria does not produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which does not produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), still particularly does not produce a β-galactosidase.
- In various embodiments, the Bacteroides is B. thetaiotaomicron and/or B. faecis. Preferably, the Bacteroides is B. thetaiotaomicron and/or B. faecis which produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- The invention encompasses the following embodiments:
- A method of preventing myocarditis in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
- A method of treating myocarditis or DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
- A method of limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
- A method of diagnosis a subject as having myocarditis or DCM, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample.
- Any of these methods, comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin. In some embodiments, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics. In some embodiments, the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants. In some embodiments, the Bacteroides is B. thetaiotaomicron and/or B. faecis.
- A composition for preventing myocarditis in a subject in need thereof, comprising a pharmaceutical agent which reduces the amount of Bacteroides sp. in a subject.
- A composition for treating myocarditis or DCM in a subject in need thereof, comprising a therapeutic agent which reduces the amount of Bacteroides sp. in a subject.
- A composition for limiting progression of myocarditis toward DCM in a subject in need thereof, comprising a pharmaceutical agent which reduces the amount of Bacteroides sp. in a subject.
- Any of these compositions, comprising an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin. In some embodiments, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics. In some embodiments, the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants. In some embodiments, the Bacteroides is B. thetaiotaomicron and/or B. faecis.
-
FIGS. 1A-R . Microbiome-dependent transition of autoimmune myocarditis to dilated cardiomyopathy. (FIG. 1A ) Survival of TCRM mice under SPF or GF conditions. (FIG. 1B ) Gross pathology of hearts from 12 week old SPF TCRM and age-matched GF TCRM mice. (FIG. 1C ) Histological analysis of hearts of 12 week old TCRM mice kept under SPF or GF conditions using hematoxylin-eosin (HE) and elastica-van Gieson (EVG) staining. (FIG. 1D ) Histopathological disease severity in TCRM mice under SPF and GF conditions. Dots represent values of individual mice; bar indicates mean disease severity. (FIG. 1E-G ) Echocardiographic parameters in TCRM mice kept under SPF or GF conditions and in transgene-negative littermate of controls with (FIG. 1E ) ejection fraction, (FIG. 1F ) fractional shortening and (FIG. 1G ) systolic left ventricular internal diameter (LVID) determined in individual mice. (FIG. 1H-J ) Co-housing of GF TCRM mice with SPF TCRM mice at the age of 4 and 8 weeks. (FIG. 1H ) Schematic representation of co-housing experiments. (FIG. 1I ) Prospective survival analysis of TCRM mice, arrowheads indicate the age of transfer to SPF conditions. (FIG. 1J ) Development of myocarditis in TCRM mice under SPF, GF and co-housing conditions; dots represent disease severity in individual mice; bar indicates mean disease severity. (FIG. 1K and L) Enumeration of heart-infiltrating cells of 12 week old TCRM mice under SPF and GF conditions or GF TCRM mice co-housed (CoH) for 4 weeks in SPF conditions with (FIG. 1K ) CD45+ cells and (FIG. 1L ) myosin-specific (Vβ8+CD4+) cells. (FIG. 1M ) IFN-γ- and (FIG. 1N ) IL-17-producing heart-infiltrating MYH6-specific CD4+ T cells (Vβ8+CD4+). (FIG. 1O-R ) Heart-infiltrating myeloid cell subsets from SPF, GF or 4 week cohoused TCRM mice analyzed by flow cytometry; dots represent values from individual mice, line indicates mean value. (FIG. 1O ) Representative tSNE plots of myeloid cells in the heart of TCRM SPF mice. Enumeration of (FIG. 1P ) inflammatory monocytes, (FIG. 1Q ) MHCIIhigh macrophages and (FIG. 1R ) MERTK+ macrophages in the heart (mean±SEM). Pooled data from 2 independent experiments n=5-6 mice (FIG. 1K , L, P-R) or three independent experiments with ≥15 mice (FIG. 1A and I), ≥12 mice (FIG. 1B , C and D), ≥6 mice (FIG. 1E-G ), ≥11 mice (FIG. 1J )≥7 mice (FIG. 1M ) per group. Representative micrographs of hearts and heart sections from one out of at least 12 mice (FIG. 1A , B). Statistical analysis was performed using Student's t test (FIG. 1D ) or one-way ANOVA with Dunnett's multiple comparison test (FIG. 1E-G , J-N and P-R) with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 2A-M . Interaction of MYH6-specific CD4+ T cells with the intestinal microbiome. (FIG. 2A ) Flow cytometric analysis of mucosal homing markers in heart-infiltrating MYH6 specific cells from SPF or GF TCRM mice. (FIG. 2B-C ) Location and proliferation of CFSE-labelled TCRM cells after adoptive transfer to Rag1−/− mice. (FIG. 2B ) Confocal microscopy analysis of colonic patches atday 3 post adoptive transfer, scale bar 100 μm (FIG. 2C ) Proliferation of MYH6-specific cells flow cytometry-based quantification of CFSE dilution in the indicated organs and at the indicated time points (mean±SEM). (FIG. 2D ) Microbiome analysis of 12 week old transgene-negative littermate controls (Tg−), SPF TCRM and GF TCRM mice co-housed at 4 weeks of age under SPF conditions. (FIG. 2E ) Principal component analysis of the fecal bacterial composition. (FIG. 2F ) Heat map of the relative abundance of the indicated bacterial classes and families in feces. (FIG. 2G-H ) MYH6-specific CD4+ T cell crossreactivity. (FIG. 2G ) In vitro proliferation of CD4+ T cells from TCRM mice after re-stimulation with MYH6, gal peptide from Bacteroides and cysteine hydrolase-derived peptide from Enterobacter determined by CFSE dilution assay. (FIG. 2H ) Cytokine production of heart- and colon-infiltrating, Vβ8-expressing CD4+ T cells from TCRM mice under SPF conditions after ex vivo re-stimulation with MYH6 peptide and Bacteroides β-gal peptide (box and whiskers show mean±interquartile range). (FIG. 2I-M ) GF TCRM mice were monocolonized with parental B. thetaiotaomicron (wild type, WT) or B. thetaiotaomicron lacking the β-galactosidase BT1626 (B. thetaiotaomicron Δβ-gal). (FIG. 2I ) Schematic representation of the experimental setting. Flow cytometric analysis of (FIG. 2J ) heart-infiltrating cells and (FIG. 2K ) MYH6-specific cytokine producing cells in the heart and colon (box and whiskers show mean±interquartile range). (FIG. 2L ) IgA bound fecal bacteria determined by bacterial flow cytometry and (FIG. 2M ) pooled data shown as mean±SEM. Pooled data from n=6 (FIG. 2A-F , J-M), n=7 (FIG. 2G ) and n=5 (FIG. 2H ) mice from at least two independent experiments. Representative histograms from two independent experiments with duplicates (FIG. 2G ). Statistical analysis was performed using Student's t test (FIG. 2A , H, J, K, M) or one-way ANOVA with Dunnett's multiple comparison test (FIG. 2F ) with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 3A-K . Impact of antibiotics treatment on lethal heart disease and immune reactivity in the TCRM model. (FIG. 3A ) Survival, (FIG. 3B ) disease severity, and (FIG. 3C ) histological analysis of hearts of TCRM mice after post-weaning oral treatment with a broad spectrum antibiotics combination comprising of Sulphadoxine, Trimethoprim and Metronidazole (S+T+M); disease severity was determined in 20 week old mice; dots represent values of individual mice; bar indicates mean disease severity. (FIG. 3D-J ) Effect of antibiotics treatment regimen on myocarditis progression in the adoptive transfer myocarditis model. (FIG. 3D ) Schematic representation of the experimental set up. (FIG. 3E ) B. thetaiotaomicron quantification in feces of mice by qPCR in the indicated treatment groups. (FIG. 3F ) Histopathological analysis of hearts from Rag1−/− mice treated with antibiotics atday 28; dots represent values of individual mice; bar indicates mean disease severity. Quantification of heart-infiltrating CD45+ cells (FIG. 3G ) and Vα8-expressing CD4+ T cells (FIG. 3H ) in Rag1−/− mice atday 28. (FIG. 3I ) Cytokine production of heart-infiltrating MYH6-specific Vβ8+CD4+ T cells (box and whiskers show mean±interquartile range). (FIG. 3J ) Heat map representation of the specific IgG responses against B. thetaiotaomicron, B distasonis, B. vulgatus and E. cloacae determined by ELISA. (FIG. 3K ) B. thetaiotaomicron-specific IgG response in sera of BALB/c mice or BALB/c mice adoptively transferred with CD4+ TCRM T cells (box and whiskers show mean±interquartile range). Pooled data from two to three independent experiments with n=7-10 (FIG. 3A-C ), n=5-13 mice (FIG. 3G-J ) and n=5-7 (FIG. 3K ). Representative sections from one out of 10 or 7 mice (FIG. 3C ). Statistical analysis was performed using Student's t test (FIG. 3K ) or Mann-Whitney test (FIG. 3B ); one-way ANOVA with Dunnett's multiple comparison test (FIG. 3E-I ) with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 4A-M . Immune reactivity against Bacteroides and cardiac myosin antigens in human myocarditis patients. (FIG. 4A ) Analysis of B. thetaiotaomicron-specific IgG antibodies in sera of myocarditis patients from the AMITIS cohort at first admission and different time points post-diagnosis compared to sera from a cohort of healthy individuals, light grey and dark grey squares indicate patients with high or low anti-B. thetaiotaomicron antibody levels respectively, cut value was determined as the mean+2SD of the healthy group; dots represent individual antibody levels, bar indicates mean value. (FIG. 4B ) Ejection fraction (EF) and (FIG. 4C ) C-reactive protein (CRP) values in patients from the AMITIS cohort at the indicated time points; individual values are shown, bar represents mean value. (FIG. 4D ) Composite clinical scores of myocarditis patients from the AMITIS cohort with low vs high IgG antibodies against B. thetaiotaomicron at the first visit (as indicated in A); dots represent individual clinical scores as the sum of positivity for anti-beta1-AR antibodies, CRP values and proportion of EF (FIG. 4E-F ) Anti-bacterial IgG antibody responses in sera of myocarditis patients and healthy controls. (FIG. 4E ) Heat map representation of specific IgG response against B. thetaiotaomicron, B. distasonis, B. vulgatus and E. cloacae in the AMITIS cohort. (FIG. 4F ) B. thetaiotaomicron-specific IgG in serum of patients of the Micro-DCM cohort at admission (individual values are shown, bar indicates mean value) and (FIG. 4G ) heat map representation of antibacterial IgG reactivity in the Micro-DCM cohort. (FIG. 4H ) Heat maps representing the binding of the MYH6614-629 peptide or the B. thetaiotaomicron β-gal11-25 peptide. The in-silico analysis was done for molecules that cover the 99% of the HLA-DQ alleles in open population. (FIG. 4I ) IFN-γ ELISPOT analysis of peripheral blood mononuclear cells of HLA-DQB1* 03 healthy volunteers (n=10) or patients from the Micro-DCM cohort (HLA-DQB1*03 n=9; other HLA n=8) after stimulation with the human-MYH6614-629 peptide or the B. thetaiotaomicron β-gal11-25 peptides (violin plots show mean±interquartile range). (FIG. 4J ) Schematic representation of fecal transplant experiment using stool from two myocarditis patients (HLA-DQB1*03:02 haplotype and B. thetaiotaomicron 16S rRNA-positive) to GF TCRM or transgene-negative (Tg−) control mice. (FIG. 4K ) B. thetaiotaomicron quantification in feces of TCRM or Tg− controls at the indicated times post fecal transplantation (box and whiskers show mean±interquartile range). (FIG. 4L-M ) Enumeration of heart-infiltrating CD45+ immune (FIG. 4L ) and CD4+ T cells (FIG. 4M ) in recipient mice at 4 weeks after fecal transplantation (mean±SEM). Statistical analysis was performed using one-way ANOVA with Dunnett's multiple comparison test (FIG. 4A-C and I) or Mann-Whitney U test (FIG. 4D ) or Student's t test (FIG. 4E-D and (L-M) with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 5A-F . Impact of microbial colonization on heart function and inflammation in TCRM mice. (FIG. 5A-C ) Echocardiographic parameters in TCRM mice under SPF, GF and co-housing conditions with (FIG. 5A ) ejection fraction, (FIG. 5B ) systolic left ventricular internal diameter (LVID) and (FIG. 5C ) fractional shortening determined in individual mice. (FIG. 5D-F ) Flow cytometric analysis of heart-infiltrating myeloid cells. Representative dot plots showing CCR2 and Ly6C (FIG. 5D ) and CD64 and MHCII expression (FIG. 5E ) shown as percentage of CD11b+ Ly6G− cells depicted for each condition. (FIG. 5F ) Enumeration of different subpopulations of myeloid cells in TCRM mice kept under the indicated conditions (mean values±SEM; n=5-8 mice per group). Statistical analysis was performed using one-way ANOVA with Dunnett's multiple comparison test with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 6A-1 . Activation TCRM CD4+ cells and interaction with the intestinal microbiome. (FIG. 6A ) Schematic representation of the experimental set up for the analysis of homing and early proliferation of CFSE-labelled TCRM cells in Rag−/− mice. (FIG. 6B-C ) Histological analysis of colonic (FIG. 6B ) and mesenteric (FIG. 6C ) lymph nodes ofRae mice 3 days after a.t. of CFSE labelled TCRM cells, scale bar=30 μm. (FIG. 6D ) Flow cytometric analysis of the proliferation patterns of TCRM cells in the colonic lamina propria and heart at the indicated time points after a.t. to Rae mice; percentages indicate CFSElow/neg MYH6-specific cells in the indicated organs. (FIG. 6E ) Cytokine profiles of heart-infiltrating and colonic Vβ8-expressing CD4+ T cells from TCRM mice at the indicated time points analyzed by flow cytometry (mean±SEM). (FIG. 6F ) Flow cytometry-based cytokine profile analysis of Vβ8-expressing CD4+ T cells from the colonic lamina propria of TCRM mice or transgene-negative littermates (Tg−). (FIG. 6G ) Production of IFN-© and IL-17 by colonic CD4+ T cells from TCRM mice or DO11.10 mice after ex-vivo re-stimulation with MYH6614-629; representative dot-plots are shown. (FIG. 6H ) Bacterial α-diversity in feces of 12 week old TCRM mice and transgene-negative littermates (Tg−) under SPF conditions. (FIG. 6I ) Relative abundance of Bacteroides and Parabacteroides in fecal samples of 12 weeks of age TCRM or Tg− mice under SPF or cohousing conditions (box and whiskers show mean±interquartile range). (n=5-13 mice per group from 2-3 independent experiments). Statistical analysis was performed using Student's t test or one-way ANOVA with Dunnett's multiple comparison test with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 7A-I . Specificity and immune interaction of Bacteroides mimic peptide-specific CD4+ T cells. (FIG. 7A ) In vitro proliferation of CD4+ T cells from TCRM mice or DO11.10 mice measured by CFSE dilution after re-stimulation with MYH6 or Bacteroides β-galactosidase peptide or ovalbumin peptide at indicated concentration (representative histograms from 1 out of 2 experiments) (FIG. 7B ) Dendritic cells loaded with MYH6 or β-gal peptide from Bacteroides at the indicated concentration were co-cultured in vitro with CD4+ T cells from TCRM mice. Maximum proliferation of CD4+ T cells was analyzed by flow cytometry and effective concentration 50 (EC50) for each peptide was determined. (FIG. 7C ) CD4+ T cells from TCRM mice were adoptively transferred into BALB/c mice. Proliferation of CD4+ T cells from TCRM mice was determined ex vivo by CFSE dilution after injection of peptide loaded dendritic cells (n=3). (FIG. 7D -FIG. 7E ) Colonization of GF TCRM mice with SPF microbiota or B. thetaiotaomicron or E. coli. Induction of IFN-© and IL-17 in Vβ8-expressing CD4+ T cells in the colonic lamina propria of TCRM mice at 8 weeks after bacterial colonization. Fold increase values were calculated using the cytokine production in GF mice as baseline (mean±SEM). (FIG. 7F ) Strategy for the generation of the β-galactosidase BT1626 deficient B. thetaiotaomicron strain. (FIG. 7G ) Colonization efficiency of WT the parental B. thetaiotaomicron (ATCC29148Δtdk) and the Δβ-gal B. thetaiotaomicron (ATCC29148Δtdk lacking theBT1626 β-galactosidase) in GF BALB/c mice (mean±SEM). (FIG. 7H-I ) Bacterial flow of fecal IgA bound bacteria in TCRM andTg − 12 weeks old mice. (FIG. 7H ) experimental dosing, (FIG. 7I ) representative dot plots left and quantification right of IgA bound bacteria; background was set using samples from Rag−/− mice (mean±SEM). (n=3-11 mice per group from 2-3 independent experiments). Statistical analysis was performed using Student's t test or one-way ANOVA with Dunnett's multiple comparison test with *, p<0.05; **, p<0.01; ***, p<0.001 -
FIGS. 8A-F . Effect of antibiotics treatment on splenic and colonic CD4+ T cells. (FIG. 8A ) Microbiota composition analysis by 16S rRNA sequencing of mice treated with a broad spectrum antibiotics cocktail containing sulphadoxine, trimethoprim and metronidazole (S+T+M) compared to untreated TCRM mice. (FIG. 8B-F ) Rag1−/− mice were adoptively transferred with 106 splenocytes from TCRM mice and treated with the indicated antibiotics. Flow cytometry-based enumeration of colonic (FIG. 8B ) CD45+ and (FIG. 8C ) Vβ8-expressing CD4+ T and (FIG. 8D ) Vβ8-expressing CD4+ T cells in spleens. (FIG. 8E ) Cytokine production of heart infiltrating Vβ8-expressing CD4+ T cells with representative dot plots. (FIG. 8F ) IFN-© and IL-17 production in colonic Vβ8-expressing CD4+ T cells. Box and whiskers show mean±interquartile range; n=6-8 mice from 2 or 3 independent experiments. Statistical analysis was performed using one-way ANOVA with Dunnett's multiple comparison test with *, p<0.05; **, p<0.01; ***, p<0.001. -
FIGS. 9A-L . B and T cell responses in myocarditis patients. (FIG. 9A-C ) Clinical parameters at admission in B. theta high or low AMITIS patients. Antibody responses in patients from the AMITIS (FIG. 9D-F ) and Micro-DCM (FIG. 9G-J ) cohorts compared to healthy individuals. IgG antibodies against E. cloacae (FIG. 9D andFIG. 9G ), E. coli (FIG. 9E andFIG. 9H ), B. distasonis (FIG. 9I ) and B. vulgatus (FIG. 9J ) were determined by ELISA and total IgG concentrations (FIG. 9F ) were determined by turbidimetry; dots represent individual patients, bar indicates mean values. (FIG. 9K ) Correlation between MYH6- or β-gal IFN-γ-producing cells in myocarditis patients of the Micro-DCM cohort and healthy individuals with the indicated HLA-DQB1 alleles; dots represent individual values. (FIG. 9L ) Graphical representation of the multiple risk factors that regulate the outcome of inflammatory cardiomyopathy. Statistical analysis was performed using Student's t test (FIG. 9G-J ) or one-way ANOVA with Dunnett's multiple comparison test with (FIG. 9D-F ) or Pearson correlation coefficient with two tailed P-value calculation.*, p<0.05. - The present invention unveils the hitherto unknown connection between the microbiome and inflammatory cardiac disease in humans. The inventors show here that commensal bacteria such as Bacteroides can serve as a source of a mimic peptide that promotes the progression of myocarditis to inflammatory cardiomyopathy. The presented invention also shows that reducing the level of or eliminating Bacteroides in situ with treatment such as antibiotics can prevent lethal heart failure.
- In at least a substantial proportion of myocarditis patients, the presence of specific bacteria in the intestinal microbiome that produce MYH6 mimic peptides and the genetic background with the particular set of HLADQB1*03 alleles determine the risk for the development of inflammatory myocarditis (
FIG. 9L ). In this scenario, cross-reactive CD4+ T cells primed in intestinal microenvironments can enter the myocardium and exacerbate the damage caused by noxious agents or processes such as infection by cardiotropic viruses or subclinical myocardial infarction (FIG. 9L ). Likewise, unleashing control over self- and cross-reactive T cells during immune checkpoint inhibitor therapy might be a reason for potentially lethal cardiac inflammation (30). Indeed, fulminant myocarditis during combination immune checkpoint blockade was found to affect patients who shared the HLADQB1*03:01 allele (31). Thus, the invention provides the rationale for extended prospective clinical trials to further dissect the connection between the microbiome-driven activation of cross-reactive CD4+ T cells, HLA-dependent genetic predisposition and inflammatory cardiomyopathy. Targeting of the microbiome of genetically predisposed myocarditis patients or susceptible patients undergoing checkpoint inhibitor treatment through antibiotics may alleviate disease severity and can therefore contribute to the prevention of the potentially lethal sequelae of inflammatory cardiomyopathy. - To treat microbiome-associated diseases or disorders, unwanted Bacteroides, typically unwanted Bacteroides producing MYH6 mimic peptides, can be eliminated by using subtractive methods such as antibiotics, phages, recombinant phages, packaged phagemids, wild-type of synthetic endolysins, wild-type of synthetic bacteriocins (such as colicins, pyocins and/or tailocins) which result in the killing of deleterious Bacteroides bacteria or by using competitive methods such as bacteria or engineered bacteria which do not produce said MYH6 mimic peptides and preferably present a competitive advantage over unwanted Bacteroides, which results in the replacement of deleterious Bacteroides bacteria by said non-deleterious bacteria or engineered bacteria. Such strategies can significantly reduce the load of Bacteroides populations.
- The invention encompasses compositions, kits and methods for eliminating a Bacteroides bacteria in situ. The compositions, kits and methods of the invention eliminate deleterious Bacteroides bacteria, in particular Bacteroides bacteria producing MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly Bacteroides bacteria producing β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), within the host microbiome by killing or reducing the growth of these Bacteroides.
- The invention encompasses the use of a vector that can transfer with high efficiency a nucleic acid, preferably a plasmid, into a bacterial population within the microbiome that allows the expression of an exogenous enzyme that will modify a gene sequence or directly kill the Bacteroides bacteria. The invention further includes methods for screening for elimination of the Bacteroides, for determining the efficiency of vectors at eliminating these Bacteroides, and for determining the effects of these vectors. Preferably, the elimination of Bacteroides bacteria in situ involves the use of antibiotics, of wild-type of synthetic bacteriocins, the use of phages, recombinant phage, packaged phagemid, wild-type of synthetic endolysins, introducing a double strand break in the DNA sequence, or any combination thereof. In an embodiment, the elimination of Bacteroides bacteria in situ involves the use of bacteria or engineered bacteria, in particular engineered Bacteroides bacteria, which do not produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which do not produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- The invention further includes methods for pre-treatment screening of patient to assess the relevancy to treat a patient based on the presence of specific bacteria in the patient intestinal microbiome that produce MYH6 mimic peptides, the genetic background of the patient with the identification of particular set of HLADQB1*03 alleles and inflammatory myocarditis symptoms of the patient. The invention further includes a pre-treatment screening of effective antibiotics, effective bacteriocins, effective phages, effective recombinant phages, effective packaged phagemids, effective endolysins, effective bacteria or engineered bacteria or any combination thereof against the targeted Bacteroides bacteria.
- Provided herein are compositions, kits and methods of treating, preventing or curing myocarditis or DCM by killing or reducing the growth of a Bacteroides sp.
- The invention encompasses the use of a vector that can transfer with high efficiency a nucleic acid, preferably a plasmid, into a bacterial population within the microbiome that allows the expression of an exogenous enzyme that will modify a gene sequence or directly kill the Bacteroides bacteria.
- The invention encompasses methods of preventing myocarditis, treating myocarditis or DCM, or limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in the subject.
- The invention encompasses methods of reducing or eliminating a Bacteroides sp. in situ.
- In one embodiment, the Bacteroides sp. is reduced or eliminated by contacting it with an antibiotic, for example, as described Gil-Cruz et al., Science 366, 881-886 (2019), which is hereby incorporated by reference.
- In one embodiment, the Bacteroides sp. is reduced or eliminated by a genetic modification that leads to the death or reduction in growth of a Bacteroides sp. bacteria, for example, as described in Bikard et al., Cell Host Microbe, Vol. 12, 177-186 (2012) and Bikard et al., Nature Biotechnology Vol. 32 (11) 1146-51 (2014).
- In one embodiment, the Bacteroides sp. is reduced or eliminated by contacting it with an endolysin, for example, as described in Gerstmans et al., Biochem Soc Trans. 2016 February; 44(1):123-8, which is hereby incorporated by reference.
- In one embodiment, the Bacteroides sp. is reduced or eliminated by contacting it with a bacteriocin (such as a colicin, a pyocin and/or a tailocin).
- In one embodiment, the Bacteroides sp. is reduced or eliminated by administering an bacteria or an engineered bacteria, in particular engineered Bacteroides sp. which do not produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which do not produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), and preferably present a competitive advantage over Bacteroides sp. to be reduced or eliminated, for example which comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar.
- By “engineered bacteria” is meant herein a bacteria which has been genetically modified, compared to the wild-type strain, in particular to express proteins of interest and/or to stop expressing unwanted proteins.
- In a particular embodiment, when said engineered bacteria comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar, said engineered bacteria are used in combination with said exogenous food source.
- In one embodiment, the method comprises contacting the unwanted bacteria with an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, wild-type or synthetic bacteriocin wild-type or synthetic endolysin or any combination thereof.
- In one embodiment, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- In one embodiment, the phage, recombinant phage, packaged phagemid encodes a wild-type or synthetic endolysin and/or a wild-type or synthetic bacteriocin
- In one embodiment, the phage, recombinant phage, packaged phagemid encodes a nuclease selected from the group consisting of CRISPR-Cas and variants, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- In one embodiment, the Bacteroides is B. thetaiotaomicron or B. faecis.
- In a preferred embodiment, the Bacteroides sp. are contacted in situ with a vector that can transfer with high efficiency a nucleic acid into the bacteria to express an exogenous enzyme (such as Cas9 or Cpf1 also known as Cas12a) in the bacteria that results in bacterial cell death or reduced growth.
- In a preferred embodiment, the exogenous enzyme can result in a genetic modification where Cas9 nuclease is used to make the desired modification. Thus, the invention contemplates introducing a double strand break in the DNA of the bacterial DNA at a specific sequence, for example with a CRISPR/Cas system, together with non-homologous end joining (NHEJ) or homologous recombination (HR) to generate the desired genetic modification. Preferably, the double strand break is generated in the presence of an editing template comprising homologous region with DNA regions located around the specific sequence located in the bacterial DNA.
- The genetic modification can be a point mutation(s), a deletion(s), insertion(s) or any combination thereof. Preferably, the genetic modification is a point mutation, an insertion or a deletion inside a coding sequence leading to a frameshift mutation or a deletion mutation. The genetic modification preferably eliminates, reduces, or increases the expression of a gene. The genetic modification can be in the translated or untranslated regions of a gene. The genetic modification can be in the promoter region of a gene or within any other region involved in gene regulation. In one embodiment, the genetic modification integrates a phage genome or exogenous DNA into the host bacterial chromosome or endogenous plasmid(s). In one embodiment, the genetic modification results in expression of an exogenous protein from an integrated exogenous DNA in the host bacterial chromosome or endogenous plasmid(s). Most preferably, the genetic modification involves either NHEJ or HR endogenous repair mechanism of the host bacteria.
- In some embodiments, the genetic modification results in the change in at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 100, 200, 500, etc. amino acids to a different amino acid. In some embodiments, the genetic modification introduces a stop codon. In some embodiments, the genetic modification is outside protein coding sequences, within RNA, or within regulatory sequences.
- In some embodiments, the genetic modification is within a β-galactosidase gene, particularly in the coding or non-coding sequence of the β-galactosidase gene, more particularly in the coding or non-coding sequence of the BT-1626 gene, more particularly in the gene encoding the protein of sequence SEQ ID NO: 1, in particular in the nucleic acid sequence SEQ ID NO: 2.
- Bacterial Elimination with Antibiotics
- Particular bacteria or groups of bacteria such as Bacteroides sp. can be eliminated by treatment with antibiotic(s). Thus, the invention encompasses methods of treating a subject with an antibiotic. Preferably, the level of Bacteroides sp. is measured before and after the treatment.
- In one embodiment, the invention encompasses a method comprising measuring the level of a Bacteroides sp., subsequently administering an antibiotic, and measuring the level of the Bacteroides sp. after the treatment.
- In some embodiments, the antibiotic is streptomycin, vancomycin, clindamycin, or metronidazole, alone or in any possible combination. In some embodiments, the antibiotic is sulphadoxine, trimethoprim, or metronidazole, alone or in any possible combination, such as described in Gil-Cruz et al., Science 366, 881-886 (2019), which is hereby incorporated by reference. In some embodiments, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- In some embodiments, the antibiotic is selected from the group consisting of penicillins such as penicillin G, penicillin K, penicillin N, penicillin O, penicillin V, methicillin, benzylpenicillin, nafcillin, oxacillin, cloxacillin, dicloxacillin, ampicillin, amoxicillin, pivampicillin, hetacillin, bacampicillin, metampicillin, talampicillin, epicillin, carbenicillin, ticarcillin, temocillin, mezlocillin, and piperacillin; cephalosporins such as cefacetrile, cefadroxil, cephalexin, cephaloglycin, cefalonium, cefaloridine, cefalotin, cefapirin, cefatrizine, cefazaflur, cefazedone, cefazolin, cefradine, cefroxadine, ceftezole, cefaclor, cefonicid, cefprozil, cefuroxime, cefuzonam, cefmetazole, cefotetan, cefoxitin, loracarbef, cefbuperazone, cefminox, cefotetan, cefoxitin, cefotiam, cefcapene, cefdaloxime, cefdinir, cefditoren, cefetamet, cefixime, cefmenoxime, cefodizime, cefotaxime, cefovecin, cefpimizole, cefpodoxime, cefteram, ceftamere, ceftibuten, ceftiofur, ceftiolene, ceftizoxime, ceftriaxone, cefoperazone, ceftazidime, latamoxef, cefclidine, cefepime, cefluprenam, cefoselis, cefozopran, cefpirome, cefquinome, flomoxef, ceftobiprole, ceftaroline, ceftolozane, cefaloram, cefaparole, cefcanel, cefedrolor, cefempidone, cefetrizole, cefivitril, cefmatilen, cefmepidium, cefoxazole, cefrotil, cefsumide, ceftioxide, cefuracetime, and nitrocefin; polymyxins such as polysporin, neosporin, polymyxin B, and polymyxin E, rifampicins such as rifampicin, rifapentine, and rifaximin; Fidaxomicin; quinolones such as cinoxacin, nalidixic acid, oxolinic acid, piromidic acid, pipemidic acid, rosoxacin, ciprofloxacin, enoxacin, fleroxacin, lomefloxacin, nadifloxacin, norfloxacin, ofloxacin, pefloxacin, rufloxacin, balofloxacin, grepafloxacin, levofloxacin, pazufloxacin, temafloxacin, tosufloxacin, clinafloxacin, gatifloxacin, gemifloxacin, moxifloxacin, sitafloxacin, trovafloxacin, prulifloxacin, delafloxacin, nemonoxacin, and zabofloxacin; sulfonamides such as sulfafurazole, sulfacetamide, sulfadiazine, sulfadimidine, sulfafurazole, sulfisomidine, sulfadoxine, sulfamethoxazole, sulfamoxole, sulfanitran, sulfadimethoxine, sulfametho-xypyridazine, sulfametoxydiazine, sulfadoxine, sulfametopyrazine, and terephtyl; macrolides such as azithromycin, clarithromycin, erythromycin, fidaxomicin, telithromycin, carbomycin A, josamycin, kitasamycin, midecamycin, oleandomycin, solithromycin, spiramycin, troleandomycin, tylosin, and roxithromycin; ketolides such as telithromycin, and cethromycin; fluoroketolides such as solithromycin; lincosamides such as lincomycin, clindamycin, and pirlimycin; tetracyclines such as demeclocycline, doxycycline, minocycline, oxytetracycline, and tetracycline; aminoglycosides such as amikacin, dibekacin, gentamicin, kanamycin, neomycin, netilmicin, sisomicin, tobramycin, paromomycin, and streptomycin; ansamycins such as geldanamycin, herbimycin, and rifaximin; carbacephems such as loracarbef; carbapenems such as ertapenem, doripenem, imipenem (or cilastatin), and meropenem; glycopeptides such as teicoplanin, vancomycin, telavancin, dalbavancin, and oritavancin; lincosamides such as clindamycin and lincomycin; lipopeptides such as daptomycin; monobactams such as aztreonam; nitrofurans such as furazolidone, and nitrofurantoin; oxazolidinones such as linezolid, posizolid, radezolid, and torezolid; teixobactin, clofazimine, dapsone, capreomycin, cycloserine, ethambutol, ethionamide, isoniazid, pyrazinamide, rifabutin, arsphenamine, chloramphenicol, fosfomycin, fusidic acid, metronidazole, mupirocin, platensimycin, quinupristin (or dalfopristin), thiamphenicol, tigecycline, tinidazole, trimethoprim, alatrofloxacin, fidaxomicin, nalidixic acid, rifampin, derivatives and combination thereof.
- In some embodiments, the Bacteroides is resistant to an antibiotic, such as β-lactams, aminoglycosides, erythromycin or tetracycline. In these cases, the gene encoding the resistance gene for that antibiotic within the Bacteroides can be modified to make the Bacteroides susceptible to the antibiotic. In a further embodiment, the modified Bacteroides is then treated with the specific antibiotic.
- The elimination of bacteria can be assessed by comparison with and without (control sample) the antibacterial treatment either in vitro or in vivo. Untreated samples can serve as control samples. The comparison is preferably performed by assessing the percentage of bacteria before and after the antibacterial treatment at at least two timepoints and determining a reduced amount of the targeted bacteria at later timepoint, for example, as described in the examples.
- Comparison in vitro can be performed by growing the bacteria in solid or liquid culture and determining the percentages or levels of a bacteria over time. The percentages or levels can be determined by routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- Comparison in vivo can be performed by collecting samples (e.g., stool or swab) over time and determining the percentages or levels of a bacteria over time. The percentages can be determined by routine diagnostic procedures employing immunodetection (e.g. ELISA), nucleic acid amplification (e.g., PCR), High Resolution Melting, and nucleic acid sequencing.
- Preferred levels of elimination of bacteria are at least 70%, 80%, 90%, 95%, 97%, 98%, 99%, 99.9%, 99.99%, and 100% of the starting levels of a bacteria.
- In some embodiments, the genetic modification is made with one or more of the following enzymes/systems:
- A) Cytosine base editors (CBE) and Adenosine base editors (ABE), for example as described in Rees et al., Nat Rev Genet. 2018 Dec.; 19(12): 770-788, which is hereby incorporated by reference.
- So far there are seven types of DNA base editors described:
-
- Cytosine Base Editor (CBE) that convert C:G into T:A (Komor, A et al. Nature 533:420-4. (2016))
- Adenine Base Editor (ABE) that convert A:T into G:C (Gaudelli, N. M. et al. Nature 551(7681) 464-471 (2017))
- Cytosine Guanine Base Editor (CGBE) that convert C:G into G:C (Chen, L et al. Precise and programmable C:G to G:C base editing in genomic DNA. Biorxiv (2020); Kurt, I et al. Nature Biotechnology 39: 41-46(2021))
- Cytosine Adenine Base Editor (CABE) that convert C:G into A:T (Zhao, D et al. Nature Biotechnology 39:35-40(2021))
- Adenine Cytosine Base Editor (ACBE) that convert A:T into C:G (WO2020181180)
- Adenine Thymine Base Editor (ATBE) that convert A:T into T:A (WO2020181202)
- Thymine Adenine Base Editor (TABE) that convert T:A into A:T (WO2020181193; WO2020181178; WO2020181195)
- Base editors differ in the base modification enzymes. CBE rely on ssDNA cytidine deaminase among which: APOBEC1, rAPOBEC1, APOBEC1 mutant or evolved version (evoAPOBEC1), and APOBEC homologs (APOBEC3A (eA3A), Anc689), Cytidine deaminase 1 (CDA1), evoCDA1, FERNY, evoFERNY.
- ABE rely on deoxyadenosine deaminase activity of a tandem fusion TadA-TadA* where TadA* is an evolved version of TadA, an E. coli tRNA adenosine deaminase enzyme, able to convert adenosine into Inosine on ssDNA. TadA* include TadA-8a-e and TadA-7.10.
- Except from base modification enzyme there has been also modifications implemented to base editor to increase editing efficacy, precision and modularity:
-
- the addition of one or two uracil DNA glycosylase inhibitor domain (UGI) to prevent base excision repair mechanism to revert base edition
- the addition of Mu-GAM that decrease insertion-deletion rate by inhibiting Non-homologous end joining mechanism in the cell (NHEJ)
- the use of nickase active Cas9 (nCas9 D10A) that, by creating nicks on the non-edited strand favors its repair and consequently the fixation of the edited base.
- the use of diverse Cas proteins from for example different organisms, mutants with different PAM motifs or different fidelity or different family (e.g. Cas12a).
- Non-limiting examples of DNA-based editor proteins include BE1, BE2, BE3, BE4, BE4-GAM, HF-BE3, Sniper-BE3, Target-AID, Target-AID-NG, ABE, EE-BE3, YE1-BE3, YE2-BE3, YEE-BE3, BE-PLUS, SaBE3, SaBE4, SaBE4-GAM, Sa(KKH)-BE3, VQR-BE3, VRER-BE3, EQR-BE3, xBE3, Cas12a-BE, Ea3A-BE3, A3A-BE3, TAM, CRISPR-X, ABE7.9, ABE7.10, ABE7.10*, xABE, ABESa, VQR-ABE, VRER-ABE, Sa(KKH)-ABE, ABE8e, SpRY-ABE, SpRY-CBE, SpG-CBE4, SpG-ABE, SpRY-CBE4, SpCas9-NG-ABE, SpCas9-NG-CBE4, enAsBE1.1, enAsBE1.2, enAsBE1.3, enAsBE1.4, AsBE1.1, AsBE1.4, CRISPR-Abest, CRISPR-Cbest, eA3A-BE3, AncBE4.
- Cytosine Guanine Base Editors (CGBE) consist of a nickase CRISPR fused to:
-
- a. A cytosine deaminase (rAPOBEC) and base excision repair proteins (e.g. rXRCC1). (Chen, L et al. Precise and programmable C:G to G:C base editing in genomic DNA. Biorxiv (2020); Chen et al. Programmable C:G to G:C genome editing with CRISPR-Cas9-directed base excision repair proteins. Nature Communications 12:1384 (2021)).
- b. A rat APOBEC1 variant (R33A) protein and an E. coli-derived uracil DNA N-glycosylase (eUNG) (Kurt, I et al. Nature Biotechnology 39: 41-46(2021)).
- Cytosine Adenine Base Editors (CABE) consist of a Cas9 nickase, a cytidine deaminase (e.g. AID), and a uracil-DNA glycosylase (Ung) (Zhao, D et al. Nature Biotechnology 39:35-40(2021)).
- ACBE include a nucleic acid programmable DNA-binding protein and an adenine oxidase (WO2020181180).
- ATBE consist of a Cas9 nickase and one or more adenosine deaminase or an oxidase domain (WO2020181202).
- TABE consist of a Cas9 nickase and an adenosine methyltransferase, a thymine alkyltransferase, or an adenosine deaminase domain (WO2020181193; WO2020181178; WO2020181195).
- Base editor molecules can also consist of two or more of the above listed editor enzymes fused to a Cas protein (e.g. combination of an ABE and CBE). These biomolecules are named dual base editors and enable the editing of two different bases (Grunewald, J et al. Nature Biotechnology 38:861-864(2020); Li, C et al. Nature Biotechnology 38:875-882(2020)).
- B) Prime editors (PE), as described in Anzalone et al. Nature 576:149-157 (2019) which is hereby incorporated by reference, consist of a nCas9 fused to a reverse transcriptase used in combination with a prime editing RNA (pegRNA; a guide RNA that includes a template region for reverse transcription).
- Prime Editing allows introduction of insertions, deletions (indels), and 12 base-to-base conversions. Prime editing relies on the ability of a reverse transcriptase (RT), fused to a Cas nickase variant, to convert RNA sequence brought by a prime editing guide RNA (pegRNA) into DNA at the nick site generated by the Cas protein. The DNA flap generated from this process is then included or not in the targeted DNA sequence.
- Prime Editing Systems Include:
-
- a Cas nickase variant such as Cas9-H840A fused to a reverse transcriptase domain such as M-MLV RT or its mutant version (M-MLV RT(D200N), M-MLV RT(D200N/L603W), M-MLV RT(D200N/L603W/T330P/T306K/W313F)
- a prime editing guide RNA (pegRNA)
- To favor editing, the prime editing system can include the expression of an additional sgRNA targeting the Cas nickase activity towards the non-edited DNA strand ideally only after the resolution of the edited strand flap by designing the sgRNA to anneal with the edited strand but not with the original strand.
- Non-limiting examples of prime editing systems include PE1, PE1-M1, PE1-M2, PE1-M3, PE1-M6, PE1-M15, PE1-M3inv, PE2, PE3, PE3b.
- C) Cas9 Retron preciSe Parallel Editing via homologY (‘CRISPEY’), a retron RNA fused to the sgRNA and expressed together with Cas9 and the retron proteins including at least the reverse transcriptase (Sharon, E. et al. Cell 175, 544-557.e16 (2018)).
- D) The SCRIBE strategy: a retron system expressed in combination with a recombinase promoting the recombination of single stranded DNA, also known as single stranded annealing proteins (SSAPs) (Farzadfard, F. & Lu, T. K. Science 346, 1256272 (2014)). Such recombinases include but are not limited to phage recombinases such as lambda red, recET, Sak, Sak4, and newly described SSAPs described in Wannier, T. M. et al. (Improved bacterial recombineering by parallelized protein discovery. Biorxiv 2020.01.14.906594 (2020) doi:10.1101/2020.01.14.906594), which is hereby incorporated by reference.
- E) The targetron system based on group II introns described in Karberg, M. et al.
Nat Biotechnol 19, 1162-7 (2001), which is hereby incorporated by reference, and which has been adapted to many bacterial species. - F) Other retron based gene targeting approaches are described in Simon et al. Nucleic Acids Res 47, 11007-11019 (2019) which is hereby incorporated by reference.
- G) CRISPR-Cas. The CRISPR system contains two distinct elements, i.e. i) an endonuclease, in this case the CRISPR associated nuclease (Cas or “CRISPR associated protein”) and ii) a guide RNA. Depending on the type of CRISPR system, the guide RNA may be in the form of a chimeric RNA which consists of the combination of a CRISPR (crRNA) bacterial RNA and a tracrRNA (trans-activating RNA CRISPR)16. The guide RNA combines the targeting specificity of the crRNA corresponding to the “spacing sequences” that serve as guides to the Cas proteins, and the conformational properties of the tracrRNA in a single transcript. When the guide RNA and the Cas protein are expressed simultaneously in the cell, the target genomic sequence can be permanently interrupted (and causing disappearance of the targeted and surrounding sequences and/or cell death, depending on the location) or modified. The modification may be guided by a repair matrix.
- The CRISPR system includes two main classes depending on the nuclease mechanism of action:
-
-
Class 1 is made of multi-subunit effector complexes and includes type I, Ill and IV -
Class 2 is made of single-unit effector modules, like Cas9 nuclease, and includes type II (II-A, II-B, II-C, II-C variant), V (V-A, V-B, V-C, V-D, V-E, V-U1, V-U2, V-U3, V-U4, V-U5) and VI (VI-A, VI-B1, VI-B2, VI-C, VI-D)
-
- The sequence of interest according to the present invention comprises a nucleic acid sequence encoding Cas protein. A variety of CRISPR enzymes are available for use as a sequence of interest on the plasmid according to the present invention. In some embodiments, the CRISPR enzyme is a Type II CRISPR enzyme, a Type II-A or Type II-B CRISPR enzyme. In another embodiment, the CRISPR enzyme is a Type I CRISPR enzyme or a Type III CRISPR enzyme. In some embodiments, the CRISPR enzyme catalyzes DNA modification. In some other embodiments, the CRISPR enzyme catalyzes RNA modification. For instance, Cas13-deaminase fusions have been used for RNA base editing thus modifying RNA (David B T Cox et al, Science, 358 (6366) p. 1019-1027, 2017 Nov. 24). In one embodiment, the CRISPR enzymes may be coupled to a guide RNA or single guide RNA (sgRNA). In certain embodiments, the guide RNA or sgRNA targets a gene selected from the group consisting of an antibiotic resistance gene, virulence protein or factor gene, toxin protein or factor gene, a bacterial receptor gene, a membrane protein gene, a structural protein gene, a secreted protein gene, a gene expressing resistance to a drug in general and a gene causing a deleterious effect to the host. Preferably, the CRISPR enzyme makes a double strand break. In some embodiments, the CRISPR enzyme makes a single strand break or nicks. In some embodiments, the CRISPR enzyme does not make any break in the DNA or RNA. In one embodiment, a Cas13-deaminase fusion is used to base edit an RNA.
- The sequence of interest may comprise a nucleic acid sequence encoding a guide RNA or sgRNA to guide the Cas protein endogenous to the targeted bacteria, alone or in combination with a Cas protein and/or a guide RNA encoded by the payload.
- Non-limiting examples of Cas proteins as part of a multi-subunit effector or as a single-unit effector include Cas1, Cas1B, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csn1 and Csx12), Cas10, Cas11 (SS), Cas12a (Cpf1), Cas12b (C2c1), Cas12c (C2c3), Cas12d (CasY), Cas12e (CasX), C2c4, C2c8, C2c5, C2c10, C2c9, Cas13a (C2c2), Cas13b (C2c6), Cas13c (C2c7), Cas13d, Csa5, Csc1, Csc2, Cse1, Cse2, Csy1, Csy2, Csy3, Csf1, Csf2, Csf3, Csf4, Csm1, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csn2, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx13, Csx1, Csx15, SdCpf1, CmtCpf1, TsCpf1, CmaCpf1, PcCpf1, ErCpf1, FbCpf1, UbcCpf1, AsCpf1, LbCpf1, homologues thereof, orthologues thereof, variants thereof, or modified versions thereof. In some embodiments, the CRISPR enzyme cleaves both strands of the target nucleic acid at the Protospacer Adjacent Motif (PAM) site.
- In various embodiments, the invention encompasses fusion proteins comprising a Cas9 (e.g., a Cas9 nickase) domain and a deaminase domain. In some embodiments, the fusion protein comprises Cas9 and a cytosine deaminase enzyme, such as APOBEC enzymes, or adenosine deaminase enzymes, such as ADAT enzymes, for example as disclosed in U.S. Patent Publ. 2015/0166980, which is hereby incorporated by reference. In one embodiment, the deaminase is an ACF1/ASE deaminase.
- In various embodiments, the APOBEC deaminase is selected from the group consisting of APOBEC1 deaminase, APOBEC2 deaminase, APOBEC3A deaminase, APOBEC3B deaminase, APOBEC3C deaminase, APOBEC3D deaminase, APOBEC3F deaminase, APOBEC3G deaminase, and APOBEC3H deaminase. In various embodiments, the fusion protein comprises a Cas9 domain, a cytosine deaminase domain, and a uracil glycosylase inhibitor (UGI) domain.
- In one embodiment, the deaminase is an adenosine deaminase that deaminate adenosine in DNA, for example as disclosed in U.S. Pat. No. 10,113,163, which is hereby incorporated by reference. In some embodiments, the fusion proteins further comprise a nuclear localization sequence (NLS), and/or an inhibitor of base repair, such as, a nuclease dead inosine specific nuclease (dISN), for example as disclosed in U.S. Pat. No. 10,113,163. In various embodiments, the invention encompasses fusion proteins comprising a catalytically impaired Cas9 endonuclease fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit, for example as described in Anzalone et al., Nature, Vol. 576, pages 149-157 (2019), which is hereby incorporated by reference.
- In a particular embodiment, the CRISPR enzyme is any Cas9 protein, for instance any naturally-occurring bacterial Cas9 as well as any variants, homologs or orthologs thereof.
- By “Cas9” is meant a protein Cas9 (also called Csn1 or Csx12) or a functional protein, peptide or polypeptide fragment thereof, i.e. capable of interacting with the guide RNA(s) and of exerting the enzymatic activity (nuclease) which allows it to perform the double-strand cleavage of the DNA of the target genome. “Cas9” can thus denote a modified protein, for example truncated to remove domains of the protein that are not essential for the predefined functions of the protein, in particular the domains that are not necessary for interaction with the gRNA(s). In some embodiments, the CAS9 is a dCas9 (dead-Cas9) or nCas9 (nickase Cas9) lacking double stranded DNA cleavage activity.
- The sequence encoding Cas9 (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cas9 protein (Fonfara et al.,
Nucleic Acids Research 42, 2577-2590 (2014); Koonin et al., Current Opinion in Microbiology 37, 67-78 (2017)). Examples of Cas9 proteins useful in the present invention include, but are not limited to, Cas9 proteins of Streptococcus pyogenes (SpCas9), Streptococcus thermophiles (St1Cas9, St3Cas9), Streptococcus mutans, Staphylococcus aureus (SaCas9), Campylobacter jejuni (CjCas9), Francisella novicida (FnCas9) and Neisseria meningitides (NmCas9). - The sequence encoding Cpf1 (Cas12a) (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cpf1 (Cas12a) protein (Koonin et al., Current Opinion in Microbiology 37, 67-78 (2017)). Examples of Cpf1 (Cas12a) proteins useful in the present invention include, but are not limited to, Cpf1 (Cas12a) proteins of Acidaminococcus sp, Lachnospiraceae bacteriu and Francisella novicida.
- The sequence encoding Cas13a (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cas13a (C2c2) protein (Abudayyeh et al., Nature 550, 280 (2017)). Examples of Cas13a (C2c2) proteins useful in the present invention include, but are not limited to, Cas13a (C2c2) proteins of Leptotrichia wadei (LwaCas13a).
- The sequence encoding Cas13d (the entire protein or a fragment thereof) as used in the context of the invention can be obtained from any known Cas13d protein (Yan et al. (2018) Mol Cell 70(2):327-339). Examples of Cas13d proteins useful in the present invention include, but are not limited to, Cas13d proteins of Eubacterium siraeum and Ruminococcus sp.
- In some embodiments, other programmable nucleases can be used. These include an engineered TALEN (Transcription Activator-Like Effector Nuclease) and variants, engineered zinc finger nuclease (ZFN) variants, natural, evolved or engineered meganuclease or recombinase variants, and any combination or hybrids of programmable nucleases. Thus, the programmable nucleases provided herein may be used to selectively modify a Bacteroides sp. DNA encoding a gene of interest.
- In some embodiments, the genetic modification is made at the RNA level. RNA base editing is based on the same principle as DNA base editing: an enzyme catalyzing the conversion of a RNA base into another must be brought close to the target base to perform its conversion locally. In one embodiment, the enzyme used for RNA editing is an adenosine deaminase from ADAR family that converts Adenosine into Inosine in dsRNA structure. Several seminal studies used this specificity for dsRNA and fused the ADAR deaminase domain (ADARDD) to an antisense oligo in order to program local RNA base editing. More recently the ability of some CRISPR-Cas systems to bind RNA molecules was repurposed into RNA editing. Using catalytically dead Cas13b enzyme (dPspCas13b) fused to a hyperactive mutant of ADAR2 deaminase domain (ADAR2DD-E488Q for REPAIRv1 and ADAR2DD-E488Q-T375G for REPAIRv2) Cox et al improved specificity and efficiency compare to previous RNA editing strategies.
- Non-limiting examples of RNA based editor proteins include REPAIRv1, REPAIRv2.
- In various embodiments, one or more of the following vectors can be used to eliminate or reduce deleterious Bacteroides sp., in particular to introduce the exogenous enzyme that results in a genetic modification or to introduce an endolysin:
-
- engineered phages,
- engineered bacteria,
- plasmid (eg, a conjugative plasmid capable of transfer into a host cell), phage, phagemid or prophage.
- Each vector may be as described above, eg, a phage capable of infecting a host cell or conjugative plasmid capable of introduction into a host cell, which can be introduced either by a phage particle (engineered or wild-type phage) via injection or by a donor bacteria via conjugation. In an example, the vectors are in combination with an antibiotic agent (eg, a beta-lactam antibiotic) and/or with any other agent.
- A bacteriophage for modifying a naturally occurring bacteria in situ comprising a nucleic acid encoding a gene editing enzyme/system for transformation of a target bacteria in a mixed bacterial population wherein said gene editing enzyme/system modifies the genome of said target bacteria, but does not lead to the death of the target bacteria.
- The invention encompasses the use of these vectors wherein the gene editing enzyme/system targets a gene within the target bacteria encoding a protein which is directly or indirectly responsible for a disease or disorder, in particular myocarditis or DCM. In a particular embodiment, the gene editing enzyme/system targets a β-galactosidase gene.
- Bacterial viruses (also called bacteriophages or phages) are small viruses displaying the ability to infect and kill bacteria while they do not affect cells from other organisms. Initially described almost a century ago by William Twort, and independently discovered shortly thereafter by Felix d′Herelle, more than 6000 different bacterial viruses have been discovered so far and described morphologically. The vast majority of these viruses are tailed while a small proportion, are polyhedral, filamentous or pleomorphic. They may be classified according to their morphology, their genetic content (DNA vs. RNA), their specific host, the place where they live (marine virus vs. other habitats), and their life cycle. As intracellular parasites of bacterial cells, phages display different life cycles within the bacterial host: lytic, lysogenic, pseudo-lysogenic, and chronic infection. Lytic phages, once their DNA injected into their host, replicate their own genome and produce new viral particles at the expense of the host. Indeed, they cause lysis of the host bacterial cell as a normal part of the final stage of their life cycles to liberate viral particles. Temperate phages can either replicate by means of the lytic life cycle and cause lysis of the host bacterium, or they can incorporate their DNA into the host bacterial DNA and become non-infectious prophages (lysogenic cycle). In some embodiments, lytic phages are used.
- Unlike classical chemically-based antibiotics that are active against a broad spectrum of bacterial species, a bacteriophage can infect and kill only a small number of different closely-related bacteria.
- The use of packaged phagemids) allows to have a defined and controlled way of killing the host. Example of packaged phagemids encoding CRISPR-Cas9 or toxins have shown promising results in killing targeted bacterial population (Bikard et al., 2012, Cell Host &
Microbe 12, 177-186; Jiang et al., 2013, Nat Biotechnol 31, 233-239; Krom et al., 2015,Nano Letters 15, 4808-4813; Bikard et al, 2014,Nat Biotech 11, Vol. 32, Citorik, R et al, 2014,Nat Biotech 11, Vol. 32). - The vector can comprise a sequence of interest under the control of a promoter.
- In one embodiment, the sequence of interest is a programmable nuclease circuit to be delivered to the targeted bacteria. This programmable nuclease circuit may be able to mediate in vivo sequence-specific elimination of bacteria that contain a target gene of interest. Some embodiments of the present disclosure relate to engineered variants of the Type II CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-associated) system of Streptococcus pyogenes. Other programmable nucleases that can be used include other CRISPR-Cas systems, engineered TALEN (Transcription Activator-Like Effector Nuclease) variants, engineered zinc finger nuclease (ZFN) variants, natural, evolved or engineered meganuclease or recombinase variants, and any combination or hybrids of programmable nucleases.
- Other sequences of interest, preferably programmable, can be added to the payload so as to be delivered to targeted bacteria.
- Preferably, the sequence of interest circuit added to the payload leads to bacteria death. The sequence of interest may comprise proteins and enzymes achieving a useful function such as modifying the metabolism of the bacteria, the composition of its environment or affecting the host.
- In a particular embodiment, the nucleic acid sequence of interest is selected from the group consisting of a Cas nuclease, a type I nuclease, a type II nuclease, a type V nuclease, a Cas9 nuclease, a Cpf1 nuclease, a Cas3 nuclease, a Cms1 nuclease, a MAD nuclease, a guide RNA, a single guide RNA (sgRNA), a CRISPR locus, a gene expressing an enzyme such as a nuclease or a kinase, a TALEN, a ZFN, a meganuclease, a recombinase, a bacterial receptor, a membrane protein, a structural protein, a secreted protein, a gene expressing resistance to an antibiotic or to a drug in general, a gene expressing a toxic protein or a toxic factor and a gene expressing a virulence protein or a virulence factor, a bacterial secretory protein or transporter, a bacterial pore or any of their combination. These proteins can also be modified or engineered to include extra features, like the addition or removal of a function (e.g. dCas9), the addition of a secretion signal to a protein not normally secreted, the addition of an exogenous peptide in a loop as non-limiting examples.
- The Bacteroides targeted by a composition of the invention can be present in vivo, in a mammalian organism, or in vitro, for example in liquid or solid culture.
- A microbiome may comprise a variety of endogenous Bacteroides species, any of which may be targeted in accordance with the present disclosure. In some embodiments, the species of targeted Bacteroides bacterial cells may depend on the type of bacteriophages being used for preparing the bacterial virus particles. For example, some bacteriophages exhibit tropism for, or preferentially target, specific host species of bacteria. Other bacteriophages do not exhibit such tropism and may be used to target a number of different genus and/or species of endogenous bacterial cells.
- Examples of Bacteroides species include, without limitation, B. acidifaciens, B. barnesiaes, B. caccae, B. caecicola, B. caecigallinarum, B. cellulosilyticus, B. cellulosolvens, B. clarus, B. coagulans, B. coprocola, B. coprophilus, B. coprosuis, B. distasonis, B. eggerthii, B. gracilis, B. faecichinchillae, B. faecis, B. finegoldii, B. fluxus, B. fragilis, B. galacturonicus, B. gallinaceumi, B. gallinarum, B. goldsteinii, B. graminisolvens, B. helcogene, B. intestinalis, B. luti, B. massiliensis, B. melaninogenicus, B. nordii, B. oleiciplenus, B. oris, B. ovatus, B. paurosaccharolyticus, B. plebeius, B. polypragmatus, B. propionicifaciens, B. putredinis, B. pyogenes, B. reticulotermitis, B. rodentium, B. salanitronis, B. salyersiae, B. sartorii, B. sedimenti, B. stercoris, B. suis, B. tectus, B. thetaiotaomicron, B. uniformis, B. vulgatus, B. xylanisolvens, and B. xylanolyticus.
- Thus, bacterial virus particles may target (e.g., specifically target) a bacterial cell from any one or more of the foregoing species of bacteria to specifically deliver the plasmid according to the invention.
- In preferred embodiments, the targeted bacterial cells are, without limitation, Bacteroides faecis, Bacteroides thetaiotaomicron, Bacteroides fragilis, Bacteroides distasonis, Bacteroides vulgatus, and Bacteroides fragilis.
- In particular embodiments, the targeted bacterial cells are, without limitation, Bacteroides sp. bacterial which produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly which produce β-galactosidase (typically of sequence SEQ ID NO: 1), more particularly which produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22).
- The bacterial virus particles are prepared from bacterial virus. The bacterial viruses can be chosen in order to be able to introduce the plasmid into the targeted bacteria.
- Bacterial viruses are preferably bacteriophages. Bacteriophages are obligate intracellular parasites that multiply inside bacteria by co-opting some or all of the host biosynthetic machinery. Phage genomes come in a variety of sizes and shapes (e.g., linear or circular). Most phages range in size from 24-200 nm in diameter. Phages contain nucleic acid (i.e., genome) and proteins, and may be enveloped by a lipid membrane. Depending upon the phage, the nucleic acid genome can be either DNA or RNA, and can exist in either circular or linear forms. The size of the phage genome varies depending upon the phage. The simplest phages have genomes that are only a few thousand nucleotides in size, while the more complex phages may contain more than 100,000 nucleotides in their genome, and in rare instances more than 1,000,000. The number and amount of individual types of protein in phage particles will vary depending upon the phage.
- A non-exhaustive listing of known Bacteroides-specific bacteria viruses is presented in the following paragraphs. Synonyms and spelling variants are indicated in parentheses. Homonyms are repeated as often as they occur (e.g., D, D, d). Unnamed phages are indicated by “NN” beside their genus and their numbers are given in parentheses.
- Bacteria of the genus Bacteroides can be infected by the following phages: crAss-phage, ad l2, Baf-44, Baf-48B, Baf-64, Bf-I, Bf-52, B40-8, FI, βI, φAl, φBrOI, φBrO2, 11, 67.1, 67.3, 68.1, mt-Bacteroides (3), Bf42, Bf71, HN-Bdellovibrio (1) and BF-41.
- In some embodiment the vectors disclosed herein may be used in combination with prebiotics. Prebiotics include, but are not limited to, amino acids, biotin, fructo-oligosaccharide, galacto-oligosaccharides, hemicelluloses (e.g., arabinoxylan, xylan, xyloglucan, and glucomannan), inulin, chitin, lactulose, mannan oligosaccharides, oligofructose-enriched inulin, gums (e.g., guar gum, gum arabic and carrageenan), oligofructose, oligodextrose, tagatose, resistant maltodextrins (e.g., resistant starch), trans-galactooligosaccharide, pectins (e.g., xylogalacturonan, citrus pectin, apple pectin, and rhamnogalacturonan-I), dietary fibers (e.g., soy fiber, sugarbeet fiber, pea fiber, corn bran, and oat fiber) and xylooligosaccharides.
- In other embodiment, the vectors, antibiotics, bacteriocins and/or endolysins disclosed herein may be used in combination with bacteria and/or engineered bacteria and/or probiotics. Probiotics include, but are not limited to lactobacilli, bifidobacteria, streptococci, enterococci, propionibacteria, saccharomycetes, lactobacilli, bifidobacteria, or proteobacteria.
- In one embodiment, said bacteria and/or engineered bacteria, in particular engineered Bacteroides sp. and/or probiotics do not produce MYH6 mimic peptides (in particular mimics of MYH6614-629 or MYH6614-628 peptides, typically of sequence SEQ ID NO: 10, 16 or 17), more particularly do not produce β-gal11-25 peptide (typically of sequence SEQ ID NO: 11, 12, 18, 19, 20, 21 or 22), and preferably present a competitive advantage over Bacteroides sp. to be reduced or eliminated.
- In a particular embodiment, said probiotic is an engineered probiotic.
- In a particular embodiment, said engineered bacteria and/or probiotics further comprise an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source, such as an exogenous sugar.
- In a particular embodiment, said engineered bacteria and/or probiotics may further be engineered to be insensitive to the vectors, phages, antibiotics, bacteriocins and/or endolysins used herein.
- In a particular embodiment, the vector, phage, antibiotic, bacteriocin and/or endolysin disclosed herein is used in combination with (i) an engineered bacteria and/or probiotic further comprising an heterologous gene or gene circuit required to import and/or metabolize a particular exogenous food source and (ii) said exogenous food source.
- The combined use of the vector disclosed herein and the herein disclosed engineered bacteria and/or probiotic and the corresponding exogenous food source typically enables said engineered bacteria and/or probiotic to fill the niche left empty by the bacteria, in particular the Bacteroides bacteria killed by the use of the vector of the invention.
- Said bacteria and/or engineered bacteria and/or probiotic can typically be administered before, simultaneously with or after said vector, phage, antibiotic, bacteriocin and/or endolysin.
- The invention encompasses methods for screening for genetic modifications in Bacteroides sp. In one embodiment, the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp., to a subject, subsequently collecting a bacterial sample from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample. The genetic modification can be the insertion of an exogenous DNA, for example, encoding an endolysin. The method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest.
- In one embodiment, the proportion of Bacteroides sp. modified vs non-modified bacteria is quantified. Preferred percentages of bacteria with the genetic modification are at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, 99.9%, 99.99%, and 100%.
- In one embodiment, the number of non-modified endogenous bacteria is quantified prior to administering a vector. The patient can also be pre-screened to determine the genetic signature of the strains the patient carries. This will allow selection of an appropriate capsid to deliver the therapeutic payload based on the genetic signature of the strains the patient carries.
- In preferred embodiments, the vector is in a pharmaceutical or veterinary composition. Preferably the vector is a bacteriophage.
- The vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- The bacterial sample can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc.
- The level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- The invention encompasses methods for determining the efficiency of a vector at inducing genetic mutations in situ. In one embodiment, the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a naturally occurring bacteria, to a subject, subsequently collecting a bacterial sample from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest and quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample.
- In preferred embodiments, the vector is in a pharmaceutical or veterinary composition. Preferably the vector is a bacteriophage.
- The vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- The bacterial sample can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc.
- The level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- The invention encompasses methods for determining the effect of a genetic mutation on bacterial growth. In one embodiment, the method comprises administering a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp., to a subject, subsequently collecting at least two sequential bacterial samples from the subject, quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest and quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest in said bacterial samples.
- In preferred embodiments, the vector is in a pharmaceutical or veterinary composition. Preferably the vector is a bacteriophage.
- The vector can be administered to the subject by any administration technique known in the art, depending on the vector and the target bacteria's expected location in or on the subject.
- The bacterial samples can be collected by any means known in the art, such as biopsy, blood draw, urine sample, stool sample, or oral/nasal swab, etc. The samples can be collected at any sequential time points. Preferably, the time between these collections is at least 3, 6, 12, 24, 48, 72, 96 hours or 7, 14, 30, 60, 120, or 365 days.
- The level of bacteria containing or not containing a genetic modification in a base of a DNA of interest can be determined by any technique known to the skilled artisan, such as routine diagnostic procedures including ELISA, PCR, High Resolution Melting, and nucleic acid sequencing.
- All of the screening methods of the invention can use any of the vectors and enzymes/systems of the invention to screen for any of the genetic modification of the invention.
- All of the screening methods of the invention can further include a step of comparing the level of bacteria containing a genetic modification in a base of a DNA of interest with the level of bacteria not containing a genetic modification the base of a DNA of interest in a bacterial sample.
- All of the screening methods of the invention can further include a step of contacting the vector with bacteria in liquid or solid culture and quantitating the level of bacteria containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample. The method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest.
- In one embodiment, the method comprises providing a vector designed to genetically modify at least one base of a DNA of interest in a gene of a Bacteroides sp. The method can further comprise contacting the vector with a Bacteroides sp. in liquid or solid culture and quantitating the level of Bacteroides sp. containing a genetic modification in said at least one base of a DNA of interest in said bacterial sample. The method can further comprise quantitating the level of bacteria not containing a genetic modification in said at least one base of a DNA of interest. The levels of bacteria containing a genetic modification in a base of a DNA of interest can be compared with the level of bacteria not containing a genetic modification the base of a DNA of interest over time in the culture. Preferably, the time between these comparisons is at least 1, 2, 3, 4, 5, 6, 12, 24, 48, 72, or 96 hours.
- The invention encompasses methods for determining the efficiency of a bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin at reducing or killing Bacteroides sp. in situ. In one embodiment, the method comprises providing a bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin, contacting the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin with a Bacteroides sp. in situ, and quantitating the level of Bacteroides sp. killed or reduced by the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin. In some embodiments, the endolysin or bacteriocin is encoded by a vector, such as a bacteriophage. The method can further comprise quantitating the level of Bacteroides not killed or reduced by the bacteria, engineered bacteria, antibiotic, bacteriocin or endolysin. The levels of bacteria can be compared over time. Preferably, the time between these comparisons is at least 1, 2, 3, 4, 5, 6, 12, 24, 48, 72, or 96 hours.
- The invention encompasses pharmaceutical and veterinary compositions comprising the vectors, bacteria, engineered bacteria, antibiotics, bacteriocins and/or endolysins of the invention.
- The invention encompasses a pharmaceutical agent which reduces the amount of Bacteroides sp. in a subject.
- The invention encompasses in situ administration of the pharmaceutical or veterinary composition to the bacteria in a subject. Any method known to the skilled artisan can be used to contact the composition with the bacterial target in situ.
- In one embodiment, the composition comprises an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
- In one embodiment, the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
- In one embodiment, the phage, recombinant phage, packaged phagemid encodes an endolysin or a bacteriocin.
- In one embodiment, the phage, recombinant phage, packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
- In one embodiment, the Bacteroides is B. thetaiotaomicron or B. faecis.
- The pharmaceutical or veterinary composition according to the invention may further comprise a pharmaceutically acceptable vehicle. A solid pharmaceutically acceptable vehicle may include one or more substances which may also act as flavoring agents, lubricants, solubilizers, suspending agents, dyes, fillers, glidants, compression aids, inert binders, sweeteners, preservatives, dyes, coatings, or tablet-disintegrating agents. Suitable solid vehicles include, for example calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidone, low melting waxes and ion exchange resins.
- The pharmaceutical or veterinary composition may be prepared as a sterile solid composition that may be suspended at the time of administration using sterile water, saline, or other appropriate sterile injectable medium. The pharmaceutical or veterinary compositions of the invention may be administered orally in the form of a sterile solution or suspension containing other solutes or suspending agents (for example, enough saline or glucose to make the solution isotonic), bile salts, acacia, gelatin, sorbitan monooleate, polysorbate 80 (oleate esters of sorbitol and its anhydrides copolymerized with ethylene oxide) and the like. The particles according to the invention can also be administered orally either in liquid or solid composition form. Compositions suitable for oral administration include solid forms, such as pills, capsules, granules, tablets, and powders, and liquid forms, such as solutions, syrups, elixirs, and suspensions. Forms useful for enteral administration include sterile solutions, emulsions, and suspensions.
- The bacterial virus particles according to the invention may be dissolved or suspended in a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fats. The liquid vehicle can contain other suitable pharmaceutical additives such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavouring agents, suspending agents, thickening agents, colours, viscosity regulators, stabilizers or osmo-regulators. Suitable examples of liquid vehicles for oral and enteral administration include water (partially containing additives as above, e.g. cellulose derivatives, preferably sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols, e.g. glycols) and their derivatives, and oils (e.g. fractionated coconut oil and arachis oil). For parenteral administration, the vehicle can also be an oily ester such as ethyl oleate and isopropyl myristate. Sterile liquid vehicles are useful in sterile liquid form compositions for enteral administration. The liquid vehicle for pressurized compositions can be a halogenated hydrocarbon or other pharmaceutically acceptable propellant.
- In some embodiments, the invention encompasses pharmaceutical or veterinary composition formulated for delayed or gradual enteric release. In preferred embodiments, formulations or pharmaceutical preparations of the invention are formulated for delivery of the vector into the distal small bowel and/or the colon. The formulation can allow the vector to pass through stomach acid and pancreatic enzymes and bile, and reach undamaged to be viable in the distal small bowel and colon.
- In some embodiments, the pharmaceutical or veterinary composition is micro-encapsulated, formed into tablets and/or placed into capsules, preferably enteric-coated capsules.
- In some embodiments, the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release, using cellulose acetate (CA) and polyethylene glycol (PEG). In some embodiments, the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using a hydroxypropylmethylcellulose (HPMC), a microcrystalline cellulose (MCC) and magnesium stearate. the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using e.g., a poly(meth)acrylate, e.g. a methacrylic acid copolymer B, a methyl methacrylate and/or a methacrylic acid ester, or a polyvinylpyrrolidone (PVP).
- In some embodiments, the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release using a release-retarding matrix material such as: an acrylic polymer, a cellulose, a wax, a fatty acid, shellac, zein, hydrogenated vegetable oil, hydrogenated castor oil, polyvinylpyrrolidone, a vinyl acetate copolymer, a vinyl alcohol copolymer, polyethylene oxide, an acrylic acid and methacrylic acid copolymer, a methyl methacrylate copolymer, an ethoxyethyl methacrylate polymer, a cyanoethyl methacrylate polymer, an aminoalkyl methacrylate copolymer, a poly(acrylic acid), a poly(methacrylic acid), a methacrylic acid alkylamide copolymer, a poly(methyl methacrylate), a poly(methacrylic acid anhydride), a methyl methacrylate polymer, a polymethacrylate, a poly(methyl methacrylate) copolymer, a polyacrylamide, an aminoalkyl methacrylate copolymer, a glycidyl methacrylate copolymer, a methyl cellulose, an ethylcellulose, a carboxymethylcellulose, a hydroxypropylmethylcellulose, a hydroxymethyl cellulose, a hydroxyethyl cellulose, a hydroxypropyl cellulose, a crosslinked sodium carboxymethylcellulose, a crosslinked hydroxypropylcellulose, a natural wax, a synthetic wax, a fatty alcohol, a fatty acid, a fatty acid ester, a fatty acid glyceride, a hydrogenated fat, a hydrocarbon wax, stearic acid, stearyl alcohol, beeswax, glycowax, castor wax, carnauba wax, a polylactic acid, polyglycolic acid, a co-polymer of lactic and glycolic acid, carboxymethyl starch, potassium methacrylate/divinylbenzene copolymer, crosslinked polyvinylpyrrolidone, polyvinylalcohols, polyvinylalcohol copolymers, polyethylene glycols, non-crosslinked polyvinylpyrrolidone, polyvinyl acetates, polyvinylacetate copolymers or any combination thereof.
- In some embodiments, the pharmaceutical or veterinary compositions are formulated for delayed or gradual enteric release as described in U.S. Pat. App. Pub. 20110218216, which describes an extended release pharmaceutical composition for oral administration, and uses a hydrophilic polymer, a hydrophobic material and a hydrophobic polymer or a mixture thereof, with a microenvironment pH modifier. The hydrophobic polymer can be ethylcellulose, cellulose acetate, cellulose propionate, cellulose butyrate, methacrylic acid-acrylic acid copolymers or a mixture thereof. The hydrophilic polymer can be polyvinylpyrrolidone, hydroxypropylcellulose, methylcellulose, hydroxypropylmethyl cellulose, polyethylene oxide, acrylic acid copolymers or a mixture thereof. The hydrophobic material can be a hydrogenated vegetable oil, hydrogenated castor oil, carnauba wax, candelilla wax, beeswax, paraffin wax, stearic acid, glyceryl behenate, cetyl alcohol, cetostearyl alcohol or and a mixture thereof. The microenvironment pH modifier can be an inorganic acid, an amino acid, an organic acid or a mixture thereof. Alternatively, the microenvironment pH modifier can be lauric acid, myristic acid, acetic acid, benzoic acid, palmitic acid, stearic acid, oxalic acid, malonic acid, succinic acid, adipic acid, sebacic acid, fumaric acid, maleic acid; glycolic acid, lactic acid, malic acid, tartaric acid, citric acid, sodium dihydrogen citrate, gluconic acid, a salicylic acid, tosylic acid, mesylic acid or malic acid or a mixture thereof.
- In some embodiments, the pharmaceutical or veterinary compositions are a powder that can be included into a tablet or a suppository. In alternative embodiments, a formulation or pharmaceutical preparation of the invention can be a ‘powder for reconstitution’ as a liquid to be drunk or otherwise administered.
- In some embodiments, the pharmaceutical or veterinary compositions can be administered in a cream, gel, lotion, liquid, feed, or aerosol spray. In some embodiments, a bacteriophage is immobilized to a solid surface using any substance known in the art and any technology known in the art, for example, but not limited to immobilization of bacteriophages onto polymeric beads using technology as outlined in U.S. Pat. No. 7,482,115, which is incorporated herein by reference. Phages may be immobilized onto appropriately sized polymeric beads so that the coated beads may be added to aerosols, creams, gels or liquids. The size of the polymeric beads may be from about 0.1 μm to 500 μm, for example 50 μm to 100 μm. The coated polymeric beads may be incorporated into animal feed, including pelleted feed and feed in any other format, incorporated into any other edible device used to present phage to the animals, added to water offered to animals in a bowl, presented to animals through water feeding systems. In some embodiments, the compositions are used for treatment of surface wounds and other surface infections using creams, gels, aerosol sprays and the like.
- In some embodiments, the pharmaceutical or veterinary compositions can be administered by inhalation, in the form of a suppository or pessary, topically (e.g., in the form of a lotion, solution, cream, ointment or dusting powder), epi- or transdermally (e.g., by use of a skin patch), orally (e.g., as a tablet, which may contain excipients such as starch or lactose), as a capsule, ovule, elixirs, solutions, or suspensions (each optionally containing flavoring, coloring agents and/or excipients), or they can be injected parenterally (e.g., intravenously, intramuscularly or subcutaneously). For parenteral administration, the compositions may be used in the form of a sterile aqueous solution which may contain other substances, for example enough salts or monosaccharides to make the solution isotonic with blood. For buccal or sublingual administration the compositions may be administered in the form of tablets or lozenges which can be formulated in a conventional manner. In a preferred embodiment, a bacteriophage and/or polypeptide of the present invention is administered topically, either as a single agent, or in combination with other antibiotic treatments, as described herein or known in the art.
- In some embodiments, the pharmaceutical or veterinary compositions can also be dermally or transdermally administered. For topical application to the skin, the pharmaceutical or veterinary composition can be combined with one or a combination of carriers, which can include but are not limited to, an aqueous liquid, an alcohol base liquid, a water soluble gel, a lotion, an ointment, a nonaqueous liquid base, a mineral oil base, a blend of mineral oil and petrolatum, lanolin, liposomes, proteins carriers such as serum albumin or gelatin, powdered cellulose carmel, and combinations thereof. A topical mode of delivery may include a smear, a spray, a bandage, a time-release patch, a liquid-absorbed wipe, and combinations thereof. The pharmaceutical or veterinary composition can be applied to a patch, wipe, bandage, etc., either directly or in a carrier(s). The patches, wipes, bandages, etc., may be damp or dry, wherein the phage and/or polypeptide (e.g., a lysin) is in a lyophilized form on the patch. The carriers of topical compositions may comprise semi-solid and gel-like vehicles that include a polymer thickener, water, preservatives, active surfactants, or emulsifiers, antioxidants, sun screens, and a solvent or mixed solvent system. U.S. Pat. No. 5,863,560 discloses a number of different carrier combinations that can aid in the exposure of skin to a medicament, and its contents are incorporated herein.
- For intranasal or administration by inhalation, the pharmaceutical or veterinary composition is conveniently delivered in the form of a dry powder inhaler or an aerosol spray presentation from a pressurized container, pump, spray, or nebuliser with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, a hydrofluoroalkane such as 1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane, carbon dioxide, or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. The pressurized container, pump, spray, or nebuliser may contain a solution or suspension of the active compound, e.g., using a mixture of ethanol and the propellant as the solvent, which may additionally contain a lubricant, e.g., sorbitan trioleate. Capsules and cartridges (made, for example, from gelatin) for use in an inhaler or insufflator may be formulated to contain a powder mix of the bacteriophage and/or polypeptide of the invention and a suitable powder base such as lactose or starch.
- For administration in the form of a suppository or pessary, the pharmaceutical or veterinary composition can be applied topically in the form of a gel, hydrogel, lotion, solution, cream, ointment, or dusting powder. Compositions of the invention may also be administered by the ocular route. For ophthalmic use, the compositions of the invention can be formulated as micronized suspensions in isotonic, pH adjusted, sterile saline, or, preferably, as solutions in isotonic, pH adjusted, sterile saline, optionally in combination with a preservative such as a benzalkonium chloride. Alternatively, they may be formulated in an ointment such as petrolatum.
- Dosages and desired drug concentrations of the pharmaceutical and veterinary composition compositions of the present invention may vary depending on the particular use. The determination of the appropriate dosage or route of administration is within the skill of an ordinary physician. Animal experiments can provide reliable guidance for the determination of effective doses in human therapy.
- For transdermal administration, the pharmaceutical or veterinary composition can be formulated into ointment, cream or gel form and appropriate penetrants or detergents could be used to facilitate permeation, such as dimethyl sulfoxide, dimethyl acetamide and dimethylformamide.
- For transmucosal administration, nasal sprays, rectal or vaginal suppositories can be used. The active compounds can be incorporated into any of the known suppository bases by methods known in the art. Examples of such bases include cocoa butter, polyethylene glycols (carbowaxes), polyethylene sorbitan monostearate, and mixtures of these with other compatible materials to modify the melting point or dissolution rate.
- The subject according to the invention is an animal, preferably a mammal, even more preferably a human. However, the term “subject” can also refer to non-human animals, in particular mammals such as dogs, cats, horses, cows, pigs, sheep, donkeys, rabbits, ferrets, gerbils, hamsters, chinchillas, rats, mice, guinea pigs and non-human primates, among others, or non-mammals such as poultry, that are in need of treatment.
- The human subject according to the invention may be a human at the prenatal stage, a new-born, a child, an infant, an adolescent or an adult at any age.
- In a particular embodiment, the subject is carrying a HLA-DQ haplotype which binds to human MYH6614629 (typically of sequence SEQ ID NO: 17); more particularly (i) a HLA-DQB1*03 variant (in particular encoded by DQB1*03:01, DQB1*03:02, DQB1*03:03, DQB1*03:04, or DQB1*03:05 alleles) and/or (ii) a HLA-DQA1*01:02 or a HLA-DQA1*05:01 variant; still particularly a HLA-DQ7 (for example encoded by DQA1*02:01/DQB1*03:01, DQA1*03:01/DQB1*03:01, DQA1*03:03/DQB1*03:01, DQA1*03:01/DQB1*03:04, DQA1*03:02/DQB1*03:04, DQA1*04:01/DQB1*03:01, DQA1*05:05/DQB1*03:01, or DQA1*06:01/DQB1*03:01 alleles), HLA-DQ8 (for example encoded by DQA1*03:01/DQB1*03:02 or DQA1*03:02/DQB1*03:02 alleles) or HLA-DQ9 (for example encoded by DQA1*02:01/DQB1*03:03 or DQA1*03:02/DQB1*03:03 alleles) haplotype.
- In a preferred embodiment, the subject has been diagnosed with, or is at risk of developing myocarditis or DCM. Diagnostic methods of determining Bacteroides sp. infection are well known by the man skilled in the art.
- In a particular embodiment, the subject has never received any treatment prior to the administration of the vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention.
- In a particular embodiment, the subject has already received at least one line of treatment, preferably several lines of treatment, prior to the administration of the vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention.
- In a particular embodiment, the subject suffers from cancer. In a more particular embodiment, the subject has already received an immune-checkpoint inhibitor treatment, more particularly a combination of anti-CTLA-4 and anti-PD-1 therapy, in particular to treat cancer.
- In a particular embodiment, the subject suffers from an auto-immune disease, in particular from sarcoidosis, rheumatoid arthritis, dermatomyositis, systemic lupus erythematosus (SLE) or giant cell myocarditis.
- In a more particular embodiment, the subject suffers from systemic lupus erythematosus (SLE). More particularly, the subject carries a mutation in the gene encoding the 3′-5′ DNA exonuclease TREX1.
- In a particular embodiment, the subject suffers from a bacterial, viral, parasital, protozoan or fungal infection, which may cause myocarditis.
- In a particular embodiment, the subject has already received an antiretroviral treatment, an antipsychotic treatment, an anticancer treatment and/or anti-parasite treatment, such as an anti-malaria treatment.
- Preferably, the treatment is administered regularly, preferably between every day and every month, more preferably between every day and every two weeks, more preferably between every day and every week, even more preferably the treatment is administered every day. In a particular embodiment, the treatment is administered several times a day, preferably 2 or 3 times a day, even more preferably 3 times a day.
- The duration of treatment with vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or with a pharmaceutical or veterinary composition according to the invention, is preferably comprised between 1 day and 20 weeks, more preferably between 1 day and 10 weeks, still more preferably between 1 day and 4 weeks, even more preferably between 1 day and 2 weeks. In a particular embodiment, the duration of the treatment is of about 1 week. Alternatively, the treatment may last as long as the infection, disorder and/or disease persists.
- The form of the pharmaceutical or veterinary compositions, the route of administration and the dose of administration of vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably of a payload according to the invention, particularly of a payload packaged into a delivery vehicle according to the invention, preferably of a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention can be adjusted by the man skilled in the art according to the type and severity of the infection (e.g. depending on the bacteria species involved in the disease, disorder and/or infection and its localization in the patient's or subject's body), and to the patient or subject, in particular its age, weight, sex, and general physical condition.
- Particularly, the amount of vectors, delivery vehicles, bacteria, engineered bacteria, antibiotics, bacteriocins or endolysins according to the invention, preferably a payload according to the invention, particularly a payload packaged into a delivery vehicle according to the invention, preferably a packaged plasmid or phagemid into a bacterial virus particle according to the invention, or of a pharmaceutical or veterinary composition according to the invention, to be administered has to be determined by standard procedure well known by those of ordinary skills in the art. Physiological data of the patient or subject (e.g. age, size, and weight) and the routes of administration have to be taken into account to determine the appropriate dosage, so as a therapeutically effective amount will be administered to the patient or subject.
- For example, the total amount of delivery vehicles, particularly a payload packaged into a delivery vehicle according to the invention, preferably a plasmid or phagemid packaged into a bacterial virus particle according to the invention, for each administration is comprised between 104 and 1015 delivery vehicles.
- As used herein, the term “vector” refers to any construct of sequences that are capable of expression of a polypeptide in a given host cell. If a vector is used then the choice of vector is dependent upon the method that will be used to transform host bacteria as is well known to those skilled in the art. Vectors can include, without limitation, plasmid vectors and recombinant phage vectors, or any other vector known in that art suitable for delivering a polypeptide of the invention to target bacteria. The skilled artisan is well aware of the genetic elements that must be present on the vector in order to successfully transform, select and propagate host cells comprising any of the isolated nucleotides or nucleic acid sequences of the invention.
- As used herein, the term «delivery vehicle» refers to any vehicle that allows the transfer of a payload into a bacterium.
- There are several types of delivery vehicle encompassed by the present invention including, without limitation, bacteriophage scaffold, virus scaffold, bacterial virus particle, chemical based delivery vehicle (e.g., cyclodextrin, calcium phosphate, cationic polymers, cationic liposomes), protein-based or peptide-based delivery vehicle, lipid-based delivery vehicle, nanoparticle-based delivery vehicles, non-chemical-based delivery vehicles (e.g., transformation, electroporation, sonoporation, optical transfection), particle-based delivery vehicles (e.g., gene gun, magnetofection, impalefection, particle bombardment, cell-penetrating peptides) or donor bacteria (conjugation).
- Any combination of delivery vehicles is also encompassed by the present invention.
- The delivery vehicle can refer to a bacteriophage derived scaffold and can be obtained from a natural, evolved or engineered capsid.
- In some embodiment, the delivery vehicle is the payload as bacteria are naturally competent to take up a payload from the environment on their own.
- In a particular embodiment, the delivery vehicle is a bacterial virus particle, in particular a packaged phagemid.
- In a particular embodiment, the delivery vehicle, in particular the bacteriophage, bacterial virus particle or packaged phagemid, comprising the vector of the invention is incapable of self-reproduction.
- In the context of the present invention, “self-reproduction” is different from “self-replication”, “self-replication” referring to the capability of replicating a nucleic acid, whereas “self-reproduction” refers to the capability of having a progeny, in particular of producing new delivery vehicles, said delivery vehicles being either produced empty or with a nucleic acid of interest packaged.
- By “delivery vehicle incapable of self-reproduction” is meant herein that at least one, several or all functional gene(s) necessary to produce said delivery vehicle is(are) absent from said delivery vehicle (and from said vector included in said delivery vehicle). In a preferred embodiment, said at least one, several or all functional gene(s) necessary to produce said delivery vehicle is(are) present in the donor cell as defined above, preferably in a plasmid or in a helper phage present in the donor cell as defined above, enabling the production of said delivery vehicle in said donor cell.
- In the context of the invention, said functional gene necessary to produce delivery vehicle may be absent through (i) the absence of the corresponding gene or (ii) the presence of the corresponding gene but in a non-functional form.
- In an embodiment, the sequence of said gene necessary to produce said delivery vehicle is absent from said delivery vehicle. In a preferred embodiment, the sequence of said gene necessary to produce said delivery vehicle has been replaced by a nucleic acid sequence of interest, in particular by a nucleic acid sequence encoding enzymes or systems for inducing genetic modifications, as defined above.
- Alternatively, said gene necessary to produce said delivery vehicle is present in said delivery vehicle in a non-functional form, for example in a mutant non-functional form, or in a non-expressible form, for example with deleted or mutated non-functional regulators. In a preferred embodiment, said gene necessary to produce said delivery vehicle is present in said delivery vehicle in a mutated form which renders it non-functional in the target cell, while remaining functional in the donor cell.
- In the context of the invention, genes necessary to produce said delivery vehicle encompass any coding or non-coding nucleic acid required for the production of said delivery vehicle.
- Examples of genes necessary to produce said delivery vehicle include genes encoding phage structural proteins; phage genes involved in the control of genetic expression; phage genes involved in transcription and/or translation regulation; phage genes involved in phage DNA replication; phage genes involved in production of phage proteins; phage genes involved in phage proteins folding; phage genes involved in phage DNA packaging; and phage genes encoding proteins involved in bacterial cell lysis.
- As used herein, the term «payload» refers to any nucleic acid sequence or amino acid sequence, or a combination of both (such as, without limitation, peptide nucleic acid or peptide-oligonucleotide conjugate) transferred into a bacterium with a delivery vehicle.
- The term «payload» may also refer to a plasmid, a vector or a cargo.
- The payload can be a phagemid or phasmid obtained from natural, evolved or engineered bacteriophage genome. The payload can also be composed only in part of phagemid or phasmid obtained from natural, evolved or engineered bacteriophage genome.
- In some embodiment, the payload is the delivery vehicle as bacteria are naturally competent to take up a payload from the environment on their own.
- As used herein, the term “nucleic acid” refers to a sequence of at least two nucleotides covalently linked together which can be single-stranded or double-stranded or contains portion of both single-stranded and double-stranded sequence. Nucleic acids of the present invention can be naturally occurring, recombinant or synthetic. The nucleic acid can be in the form of a circular sequence or a linear sequence or a combination of both forms. The nucleic acid can be DNA, both genomic or cDNA, or RNA or a combination of both. The nucleic acid may contain any combination of deoxyribonucleotides and ribonucleotides, and any combination of bases, including uracil, adenine, thymine, cytosine, guanine, inosine, xanthine, hypoxanthine, isocytosine, 5-hydroxymethylcytosine and isoguanine. Other examples of modified bases that can be used in the present invention are detailed in Chemical Reviews 2016, 116 (20) 12655-12687. The term “nucleic acid” also encompasses any nucleic acid analogs which may contain other backbones comprising, without limitation, phosphoramide, phosphorothioate, phosphorodithioate, O-methylphosphoroamidite linkage and/or deoxyribonucleotides and ribonucleotides nucleic acids. Any combination of the above features of a nucleic acid is also encompassed by the present invention.
- As used herein the term “phagemid” or “phasmid” are equivalent and refer to a recombinant DNA vector comprising at least one sequence of a bacteriophage genome and which is preferably not able of producing progeny, more particularly a vector that derives from both a plasmid and a bacteriophage genome. A phagemid of the disclosure comprises a phage packaging site and optionally an origin of replication (ori), in particular a bacterial and/or phage origin of replication. In one embodiment, the phagemid does not comprise a bacterial origin of replication and thus cannot replicate by itself once injected into a bacterium. Alternatively, the phagemid comprises a plasmid origin of replication, in particular a bacterial and/or phage origin of replication.
- As used herein, the term “packaged phagemid” refers to a phagemid which is encapsidated in a bacteriophage scaffold, bacterial virus particle or capsid. Particularly, it refers to a bacteriophage scaffold, bacterial virus particle or capsid devoid of a bacteriophage genome. The packaged phagemid may be produced with a helper phage strategy, well known from the man skilled in the art. The helper phage comprises all the genes coding for the structural and functional proteins that are indispensable for the phagemid according to the invention to be encapsidated. The packaged phagemid may be produced with a satellite virus strategy, also known from the man skilled in the art. Satellite virus are subviral agent and are composed of nucleic acid that depends on the co-infection of a host cell with a helper virus for all the morphogenetic functions, whereas for all its episomal functions (integration and immunity, multicopy plasmid replication) the satellite is completely autonomous from the helper. In one embodiment, the satellite genes can encode proteins that promote capsid size reduction of the helper phage, as described for the P4 Sid protein that controls the P2 capsid size to fit its smaller genome.
- As used herein, the term “peptide” refers both to a short chain of at least 2 amino acids linked between each other and to a part of, a subset of, or a fragment of a protein which part, subset or fragment being not expressed independently from the rest of the protein. In some instances, a peptide is a protein. In some other instances, a peptide is not a protein and peptide only refers to a part, a subset or a fragment of a protein. Preferably, the peptide is from 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 amino acids to 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 40, 50, 100, 200 amino acids in size.
- MYH6-specific TCR transgenic mice (TCR-M) on the BALB/c background have been previously described (8). TCRM mice were maintained as heterozygous and transgene-negative littermates were used as controls. TCRM mice were re-derived to germ-free conditions by axenic 2-cell stage embryo transfer into pseudopregnant germ-free recipient females at the Clean Mouse Facility, University of Berne, Switzerland, and bred and maintained under germ-free conditions in flexible film isolators. Germ-free status was routinely monitored by culture-dependent (aerobic and anerobic) and -independent (Sytox DNA stain of cecal contents) methods and were additionally confirmed pathogen-free. In the indicated experiments, germ-free TCRM mice were transferred at 4 or 8 weeks of age to the SPF facility in the Institute of Immunobiology, Kantonsspital St. Gallen and co-housed with SPF TCRM mice and transgene-negative littermates. Rag1−/− and DO11.10 mice were obtained from the Jackson laboratories. Homozygous OVA-TCR transgenic mice (DO11.10) were mated with SPF BALB/c Thy1.1 mice from the TCRM colony of the St. Gallen facility and offspring were used for analysis. Rag1−/− mice were co-housed with
TCRM mice 3 weeks before set-up of the breeding pairs. All mice were on the BALB/c genetic background and were maintained in individually ventilated cages. Experiments were performed in accordance with federal and cantonal guidelines (Tierschutzgesetz) under permission numbers SG15/06, SG16/07, SG17/07 and BE55/14 following review and approval by the respective Cantonal Veterinary Offices (St. Gallen and Berne, Switzerland). - Serum samples from patients enrolled in two different clinical studies were analyzed: (i) “Cardiac β1-adrenoceptor autoantibodies in human heart disease: rationale and design of the Etiology, Titre-Course, and Survival” (ETiCS, AMITIS arm) (27), EC permission No 186/07 (Table 2) and (ii) “Dissecting the role of cross-reactive antimicrobial T and B cell responses in myocarditis and heart failure—an exploratory research project Micro-DCM”, EC permission No 2017-01853 (Table 3 and Table 4).
-
TABLE 2 Demographic details and HLA-DQB1* alleles of ETiCS-AMITIS myocarditis patients and healthy control group1 Diagnosis by Patient No. Age (years)2 Sex3 HLA-DQB1* EMB 41 21.3 Male 03:01 03:01 Yes 2 32.8 Male 03:01 06:02 Yes 3 58.3 Female 03:02 06:03 Yes 4 54.4 Female 02:02 06:04 Yes 5 52.8 Male 03:01 03:01 Yes 6 50.1 Male 03:02 06:02 Yes 7 35.2 Male 03:02 03:02 Yes 8 31.1 Male 02:01 03:01 Yes 9 64.7 Male 02:01 03:02 Yes 10 42.0 Male 03:02 04:02 Yes 11 24.1 Male 03:02 03:03 Yes 12 21.4 Male 02:02 03:01 Yes 13 23.8 Male 03:01 06:04 Yes 14 46.1 Male 03:01 03:03 Yes 15 20.1 Male 03:03 06:02 Yes 16 28.9 Male 05:01 06:02 Yes 17 39.5 Male 02:02 05:03 Yes 18 21.1 Male 04:02 06:02 Yes 19 25.3 Male 03:01 06:84 Yes 20 27.7 Male 02:02 04:02 Yes 21 18.5 Male 05:01 06:03 Yes 22 69.0 Male 03:01 05:03 Yes 23 18.6 Male 03:01 03:01 Yes 24 53.7 Male n.d. 5 n.d. 5 Yes 25 32.0 Male n.d. 5 n.d. 5 Yes 1Healthy controls sera were obtained at the base line visit of the interventional study registered as ISRCTN18360696. Female to male ratio of 1:0.38 (N = 18) and mean age of 35 ± 8.3 years. 2Mean age of is 36.5 ± 15.5 years in ETiCS/AMITIS. 3Female to male ratio of 1:11.5 in ETiCS/AMITIS. 4EMB: Endomyocardial biopsy. 5 n.d. non-determined. -
TABLE 3 Demographic details, HLA-DQB1* alleles and diagnosis of Micro-DCM patients. Patient No. Age (years)1 Sex2 HLA-DQB1* Diagnosis 31 69.9 Female 03:01 03:01 Heart failure 2 23.6 Male 02:01 03:01 Myocarditis 3 19.2 Male 03:02 06:03 Myocarditis 4 25.8 Male 03:01 03:02 Myocarditis 5 23.9 Female 02:02 06:03 Myocarditis 6 40.9 Female 05:01 06:03 Myocarditis 7 37.0 Male 05:01 06:03 Myocarditis 8 54.9 Male 02:01 04:02 Heart failure 9 63.5 Male 02:01 02:02 Myocarditis 10 65.0 Male 03:02 06:02 Myocarditis 11 42.8 Male 02:01 03:03 Myocarditis 12 47.7 Male 02:01 06:03 Heart failure 13 20.0 Male 03:03 06:02 Myocarditis 14 39.1 Female 03:03 05:03 Myocarditis 15 26.0 Male 04:02 05:01 Myocarditis 16 51.2 Male 02:02 05:03 Heart failure 17 30.5 Male 03:01 05:03 Myocarditis 1Mean age in myocarditis patients is 35.8 ± 15.2 and 55.9 ± 9.7 years for heart failure patients. 2Female to male ratio for myocarditis patients of 1:3.3 and 1:3 for heart failure patients. 3Myocarditis: patients with typical late enhancement pattern in cardiac MRI and exclusion of coronary artery disease by angiography or computed tomography. Heart failure: patients with left ventricular ejection fraction ≤40%, no obvious mechanism underlying left ventricular dysfunction and no significant coronary artery by invasive angiography (no stenosis >50% in any main vessel). -
TABLE 4 Demographic details and HLA-DQB1* alleles of healthy volunteers. Volunteer No. Age (years)1 Sex2 HLA-DQB1* 1 31.5 Female 02:01 03:03 2 40.3 Male 03:01 05:03 3 33.2 Male 03:01 05:01 4 55.4 Male 03:01 03:01 5 27.7 Female 06:04 06:02 6 35.1 Male 03:01 05:01 7 26.7 Female 02:02 03:19 8 31.8 Male 03:01 05:01 9 31.1 Male 03:03 05:02 10 41.2 Female 03:05 03:01 11 46.5 Female 03:02 05:02 12 32.8 Female 02:02 06:04 1Mean age 36.1 ± 8.3 years. 2Female to male ratio 1:1. - All study Participants provided written informed consent in accordance with the Declaration of Helsinki and the International Conference on Harmonization Guidelines for Good Clinical Practice. All regulations were followed according to the German or Swiss authorities and according to the clinical protocols. The retrospective part of this study (AMITIS cohort) is a secondary investigation using patient samples collected from existing clinical trials. Therefore, the sample sizes in this report were determined by the original clinical trial designs and sample availability; no additional inclusion/exclusion criteria were applied. The sample size calculation for the Micro-DCM study has not been done because of the exploratory nature of the study design.
- Histological analysis was performed as previously described (8). Briefly, hearts were fixed in 4% formaldehyde (Formafix) for at least 12 h and embedded in paraffin. Histopathological changes were evaluated following hematoxylin/eosin (HE) and Elastica van Gieson (EVG) staining. Myocarditis severity was evaluated using a semiquantitative scoring system: 0, no inflammation; 1, <100 inflammatory cells involved, small inflammatory lesions; 2, >100 inflammatory cells involved, larger inflammatory lesions; 3, >10% of the heart section involved in inflammation; 4, >30% of the heart section involved in inflammation; 5, >30% of the heart section involved in inflammation with extensive fibrosis and dilation of ventricle. Images from heart sections were acquired using a Leica DMRA microscope.
- For the generation of single-cell suspensions, spleens and lymph nodes were collected in BSS and were mechanically disrupted on a metallic grid. To confirm the presence of heart-infiltrating cells, mice received 2 μg of anti-CD45.2 BV510 or anti-Thy 1.1 PE i.v. 5 min before being euthanized in the indicated experiments. Euthanized animals were perfused with 20 ml PBS and small heart tissue pieces were placed into a 6-well dish filled with RPMI 1640 medium containing 2% FCS, 20 mM HEPES (Lonza), 1 mg/ml collagenase D (Sigma) and 25 μg/ml DNasel (Applichem) and incubated at 37° C. under continuous stirring. The remaining tissue pieces were mechanically disrupted and mononuclear cells were purified by centrifugation (25 min at 800×g, 4° C.) on a 30-70% Percoll gradient (GE Healthcare). For isolation of colonic lamina propria cells, colons were flushed and tissue was harvested and incubated three times for 15 min at room temperature under constant agitation with BSS containing 5% FCS (Lonza), 5 mM EDTA (Sigma), 10 mM HEPES (Sigma) and 1 mM DTT in order to dissociate the epithelial layer. The tissue was subsequently washed with BSS containing 10 mM HEPES and digested three times for 20 min at 37° C. with 120 μg/ml collagenase P (Roche), 25 μg/ml DNAse I (Applichem) and 5 μg/ml Dispase I (Roche) in RPMI-1640; after obtaining a single cell suspension, cells were purified by a 30-70% Percoll gradient. Single-cell suspensions were first stained with the fixable viability dye Zombie Aqua (Biolegend) and incubated for 30 min on ice; after washing, cells were incubated for 20 min at 4° C. in PBS containing 2% FCS and 10 mM EDTA with fluorochrome-labeled antibodies (Table 5). Cells were acquired with a BD LSRFortessa (BD Biosciences) and analyzed using FlowJo software (Treestar Inc) following stablished guidelines (32).
-
TABLE 5 Antibodies used in this study Clone Reagent Conjugate Dilution Source IA8 anti-Ly6G PE 1:200 Pharmigen AL-21 anti-Ly6C PerCP 1:100 Biolegend 145-2C11 anti-CD3e PerCP 1:100 Biolegend 30-F11 anti-CD45 APC-Cy7 1:100 Biolegend 30-F11 anti-CD45 PE 1:100 Biolegend IM7 anti-CD44 APC-Cy7 1:100 Biolegend MEL-14 anti-CD62L BV-421 1:100 Biolegend M1/70 anti-CD11b BV-711 1:100 Biolegend FJK16S anti-FoxP3 PeCy7 1:100 eBioscience B20.1 anti-Vα2 APC 1:100 BDBioscience MR5-2 anti-Vβ8 FITC 1:100 BDBioscience RM4-5 anti-CD4 BV-605 1:100 Biolegend XMG-1.2 anti-IFN © APC 1:100 Biolegend TC11- anti-IL17 PE 1:100 Biolegend 18H10.1 KJ1-26 anti-TCR D011.10 FITC 1:100 Biolegend 2G9 anti-IA/IE FITC 1:100 BDBioscience SA011F11 anti-CX3CR1 APC- 1:100 Biolegend Fire750 2B10C42 anti-MERTK APC 1:100 Biolegend 104 anti-CD45.2 BV-605 1:100 Biolegend SA203G11 anti-CCR2 BV-421 1:100 Biolegend X54-5/7.1 anti-CD64 PeCy7 1:250 Biolegend DATK32 anti-α4β7 APC 1:100 Biolegend 29-2L17 anti-CCR6 PE 1:100 Biolegend 11-44-2 anti-IgA PE 1:50 eBiosciences - Murine colonic tissue and lymph nodes were fixed overnight at 4° C. in freshly prepared 4% paraformaldehyde (Merck Millipore) under agitation. Tissues were embedded and oriented in 4% low melting agarose (Invitrogen) in PBS and serially sectioned with a vibratome (Leica VT-1200). 30-40 μm thick sections were blocked in PBS containing 10% FCS, 1 mg/ml anti-Fcγ receptor (BD Biosciences) and 0.1% Triton X-100 (Sigma). Tissues were incubated overnight at 4° C. with the following antibodies: anti-mouse CD4, anti-mouse E-Cadherin (Biolegend). Microscopy was performed using a confocal microscope (LSM-710, Carl Zeiss). Microscopy data were recorded with ZEN 2010 software (Carl Zeiss, Inc.) and processed in Imaris Versions 7 (Bitplane).
- For assessment of ex vivo production of IFN-γ and IL-17, 106 lymphocytes were incubated for 3.5 h at 37° C. in 96-well round-bottom plates in 200 μL of RPMI per 5% FCS supplemented with 10 μg/mL brefeldin A (Sigma). Cells were stimulated with 0.25 μg of the different peptides (see below), or phorbol myristate acetate (50 ng/ml; Sigma)/ionomycin (500 ng/ml; Sigma) (PMA/I) as positive control, or were left untreated. After surface molecule labelling, cells were stained with the fixable viability dye Zombie Aqua (Biolegend). Cells were fixed with cytofix-cytoperm (BD Biosciences) for 20 min. Fixed cells were incubated at 4° C. for 40 min with permeabilization buffer (2% FCS, 0.5% saponin in PBS) containing antibodies to CD3, CD4, IFN-γ and IL-17A. Samples were measured using a BD LSR Fortessa (BD Biosciences). Data were analyzed using FlowJo software (Tree Star, Inc.).
- Generation of B. thetaiotaomicron ΔBT1626 Mutant Strain
- Deletion of the B. thetaiotaomicron β-galactosidase (BT1626) was done using a counterselection system described in (33). A strain with a clean deletion of tdk (BT2275) in the VPI-5482 background, resistant to the toxic nucleotide analog 5-fluoro-2′-deoxyuridine (5FUdR) was used as a starting point.
Genomic regions 1 kb upstream and downstream of BT1626, respectively, were amplified via colony per, with primers including overhangs for Gibson assembly into the plasmid pExchange-tdk (33), digested with the enzymes XbaI and SaII. All primers are listed in Table 6. -
TABLE 6 Primers used for the deletion of B. thetaiotaomicron BT1626 Name Sequence Purpose Bt_bgal_ko_1 AAG ATA ACA TTC GAG TOG Forward primer 1 kb upstream ACT ATT TCG GCA ATA ATA of BT1626, with Gibson CGC TGA AAG (SEQ ID NO: overhang homologous to 3) pExchange-tdk. Bt_bgal_ko_2 CAA ATG CAA TCA GTT TCA Reverse primer immediately GGC ATT ATT ATA TTG TTT upstream of BT1626, with TTT GGT GAC TGG T (SEQ Gibson overhang homologous ID NO: 4) to the downstream region of BT1626. Bt_bgal_ko_3 CCT GAA ACT GAT TGC ATT Forward primer immediately TGG (SEQ ID NO: 5) downstream of BT1626 Bt_bgal_ko_4 GCG GTG GCG GCC GCT Reverse primer 1 kb CTA GAA GTC GTT TGT TGT downstream of BT1626, with TTT CTT CG (SEQ ID NO: 6) Gibson overhang homologous to pExchange-tdk. 1626_250us_f CAC ATA GTA GTT TTC TTT Control primer 250 bp CGT C (SEQ ID NO: 7) upstream of BT1626. 1626_264inside_r GCG AAA CTG ACG GAT Control primer 264 bp inside CAG (SEQ ID NO: 8) the BT1626 coding sequence. 1626_250ds_r TCG GAC GAT AAT GCG Control primer 250 bp ACT T (SEQ ID NO: 9) downstream of BT1626. - The plasmid was then transformed into the conjugative E. coli strain SM10 and conjugated into the B. thetaiotaomicron Δtdk. The resulting colonies are resistant to erythromycin and have the complete plasmid inserted into the chromosome resulting from a single homologous recombination event. Counterselection on plates containing 200 μg/ml 5FUdR led to a second recombination event, and either to a reversion to the wild type state or to the deletion of BT1626. Successful deletion was tested using PCR with control primers, and clones with a clean deletion of the gene were further used (
FIG. 7F and Table 6). All plates used for generating mutants were Brain Heart Infusion (Oxoid), supplemented with 2% agar (Sigma), 10% defibrinated sheep blood (Sigma), 1 μg/ml Hemin (Sigma). 200 μg/ml FUdR (Sigma), 25 μg/ml erythromycin (Sigma), and 200 μg/ml gentamicin (Sigma) were added before pouring the plates when needed. - The different Bacteroides thetaiotaomicron strains (ATCC29148, ATCC29148Δtdk or ATCC29148ΔtdkΔBT1626) were grown anaerobically overnight at 37° C. in Brain Heart Infusion (BHI, CM1135, Oxoid) with the addition of hemin (1 μg/ml, Sigma) and menadione (0.5 μg/ml, Sigma). E. coli BL21(DE3) was grown in LB medium at 37° C. Overnight broth culture of B. thetaiotaomicron or E. coli was washed once in PBS and gavaged intra-gastrically (500 μl/mouse corresponding to 109 bacteria/mouse) to germ-free mice (at least 3-4 week old). Germ-free and monocolonized animals were kept in flexible film isolators until the end of the experiments and monitored regularly for excluding any contaminations using Sytox staining on fecal pellets for GF animals, Gram staining and 16S rRNA sequencing on fecal pellets for monocolonized animals (34). The effects of mono-colonization in TCRM mice were analyzed 8 weeks post colonization.
- Spleens were collected from TCRM mice and disrupted on a 70-μm cell strainer. Red blood cells were lysed by osmotic shock and 106 splenocytes were injected intravenously in 4 wk mice in the lateral tail vein of Rag1−/− mice. Seven days after adoptive transfer, mice were bled to confirm CD4+ T cell expansion. Disease activity scores and analysis of T cell activation in the heart and colonic lamina propria were performed at
day 28 post adoptive transfer. - Mice were treated for 4 or 16 weeks with the following antibiotics in the drinking water (i) a broad spectrum antibiotic mixture; Sulfadoxine 400 mg/l and
Trimethoprim 80 mg/l (Borgal; Veterinaria) or (ii)Streptomycin 200 mg/l (Sigma-Aldrich) to reduce the proportion of Bacteroidetes (35) or iii)Vancomycin 100 mg/l. Metronidazole (Sigma-Aldrich) was administered by gavage at a dose of 2 mg every 3 days to Sulfadoxine and Trimethoprim-treated mice or a separate group of mice for exclusive Metronidazole treatment. - Spleen cells were labelled with 10 μl of 5 mM carboxyfluorescein succinimidyl ester (CSFE, dissolved in DMSO) (Molecular Probes) in 10 ml of PBS for 10 min at 37° C. The staining reaction was stopped with 1 mL FCS (Lonza), followed by washing with PBS. A total of 2×105 splenocytes/well were seeded in 96-well round-bottom plate and MYH6614-629 (RSLKLMATLFSTYASADR, SEQ ID NO: 10) or Bacteroides β-galactosidase11-25 (TFLILMAALTATFASAQK, SEQ ID NO: 11), Enterobacter cysteine hydrolase10-27 (LLIGMMSTFSTYASAQET, SEQ ID NO: 12) or OVA323-339 (ISQAVHAAHAEINEAGR, SEQ ID NO: 13) peptide was added at the indicated concentrations. CSFE dilution was assessed by flow cytometry after 3 days of incubation at 37° C. All peptides were synthetized by Genscript with >80% of purity.
- Spleen cells were labelled with CFSE as described above and 5×106 cells were i.v. transferred in to Rag−/− mice. Mice were culled at
days - High-binding 96-well polystyrene plates (Corning) were coated with 107 CFU of the following bacterial strains B. theta ATCC29148, B. distasionis, B. vulgatus, E. cloacae ATCC 13047 or E. coli K12 in 0.1 M carbonate-bicarbonate buffer, pH 9.5. Plates were incubated for 1 h at 37° C. and then overnight at 4° C. Before use, plates were washed three times with PBS containing 0.05% Tween-20 (PBS-T) (Sigma-Aldrich). Non-specific binding was blocked with 5% non-fat dry milk diluted in PBS (PBS-M) for 1 h at 37° C. After washing, sera were diluted 1:20 in PBS-M and two fold serial dilutions were added to the wells. Plates were incubated for 1 h at 37° C., followed by four washes with PBS-T and then incubated for 1 h of at 37° C. with peroxidase-conjugated rabbit anti-human IgG (1:2500) or rabbit-anti-mouse IgG (1:1000) antibody (in PBS-M, Jackson Immuno Research). After four washes with PBS-T, ortho-phenylenediamine (0.5 mg/ml; Sigma) in 0.1 M citrate buffer, pH 5.6, containing 0.08% H2O2 was used to develop the reaction. Optical density was measured at 492 nm using an automated ELISA plate reader (Tecan). Absorbance values at 1:160 dilutions are presented in the figures. In human samples high and low responders to B. thetaiotaomicron were high responders were defined as patients that had an IgG at diagnosis of A >mean of healthy controls +2 SD.
- Fecal-bound IgA was assessed using bacterial flow cytometry for IgA-bound bacteria performed as previously described (36). In brief, fecal pellets were homogenized in 1 ml of staining buffer (PBS+1% (w/v) bovine serum albumin (BSA, Sigma). Tubes were centrifuged at 400 g for 5 minutes to pellet large debris and the supernatant was filtered through a sterile 70 μm cell strainer and transferred to a new tube. Samples were centrifuged at 8000×g for 5 minutes to pellet bacteria and pellets were re-suspended in PBS with SYTO BC (1:1000, Life Technologies) for 15 minutes on ice, Samples were washed with PBS and stained in 100 μl staining buffer containing PE-conjugated anti-mouse IgA (1:75) for 30 minutes on ice. Samples were washed 3 times with PBS and re-suspended for flow cytometry analysis in a FACS CANTO (BD). Bacteria were gated based on forward and side scatter and the gating strategy was verified by SYTO-BC staining. Fecal samples from Rag1−/− mice were used as negative controls to define the threshold for IgA− and IgA+ populations. Analysis of IgA binding was done using FlowJo (v10.1).
- For microbial composition analysis, the feces from TCRM SPF mice, Tg− littermates and TCRM GF co-housed with TCRM SPF and Tg− littermates were used. Microbial composition was assessed by a 16S high-throughput amplicon analysis as described previously (37). The 163 rRNA gene segments spanning the variable V5 and V6 regions were amplified from DNA from the samples, using a multiplex approach with the barcoded
forward fusion primer 5′-CCATCTCATCCCTGCGTGTCTCCGACTCAG (SEQ ID NO:14) BARCODE ATTAGATACCCYGGTAGTCC-3′ (SEQ ID NO: 23) in combination with thereverse fusion primer 5′-CCTCTCTATGGGCAGTCGGTGATACGAGCTGACGACARCCATG-3′ (SEQ ID NO: 15). The PCR-amplified 16S V5-V6 amplicons were purified and prepared for sequencing on the Ion torrent PGM system according to the manufacturers instructions (Life Technologies). Samples with over 10′000 reads were accepted for analysis. Data analysis was performed using the QIIME pipeline version 1.8.0 (38). Operational taxonomic units were picked using UCLUST with a 97% sequence identity threshold, followed by taxonomy assignment using either the latest Greengenes database. For estimation of species diversity within samples (α-diversity), the Shannon index was calculated using the R phyloseq package after rarefaction to even depth to the sample with the lowest number of sequences (34). - Fecal Bacterial DNA Extraction and Quantitative B. thetaiotaomicron PCR
- Bacterial genomic DNA was extracted using a commercial kit (QIAamp DNA Stool Mini Kit) with minor modifications. Glass beads (Sigma, G4649-10G) were used for an optimal disruption of fecal content and an additional extraction step was introduced after incubation of bacteria with a lysis buffer (20 mg/ml lysozyme—Sigma 62970-5G-F, 2 mM EDTA, 1.2% Triton, 20 mM Tris-HCI). Extracted genomic DNA was used as a template for B. thetaiotaomicron PCR amplification and quantification using a real-time PCR machine (Quant Studio 5) and SYBR Green PCR technology (Applied Biosystems, Thermo Fisher Scientific). B. thetaiotaomicron was quantified using species specific primers that have been established and validated before (39) (atcgcaaaaataagatgggcaaa and cacaacagccatagcgttcca) with a cycling protocol of denaturation at 95° C. (2 min), 40 cycles of 95° C. (20 sec), 58° C. (20 sec) and 72° C. (30 sec), Each 17 μl qPCR mixture consisted of 8.5 μl of 2× SYBR Green MasterMix (Applied Biosystems, Thermo Fisher Scientific), 1.7 μl of BSA (100 μg/ml), 0.6 μl of each primer (10 μM), 3.60 μl PCR-grade water and 2 μl of extracted genomic DNA. DNA extracted from B. theta ATCC29148 was used to construct a standard curve with 10-fold dilutions of DNA templates of known concentrations. Concentrations of DNA used in the standard curves ranged from 33.75 ng/μl equivalent to 107 bacteria to 0.035 pg/μl equivalent to 101 bacteria. The PCR amplifications were performed in duplicates and the detection limit was 0.34 pg/μl (102 bacteria). Bacterial qPCR signals were normalized to 1 μg of total bacterial DNA.
- Proteins encoded in cultured and uncultured bacteria of the NCBI microbial protein database were searched for 15 amino acid sequences similar to MYH6614-629 with BLAST p2.3.1 (blast.ncbi.nlm.nih.gov). Predicted binding of the candidate bacterial peptides to the BALB/c mouse IAd MHC class II was evaluated using the immune epitope database and analysis resource IEDB from the National Institute of Allergy and Infectious diseases online prediction tool (tools.iedb.org/mhcii/). The binding threshold was set at 10, and peptides with percentile ranks lower than the threshold value were predicted to be good MHC II haplotype binders (Table 1).
-
TABLE 1 Sequences and properties of microbial peptide mimics of MYH6614-629. Protein Iad Murine length binding MYH6 Accession (amino Amino acid (percent- identity Description Organism number acids) sequence2 ile)3 (%) Myosin heavy Mus musculus NP_034986.1 1938 SLKLMATLFSTYASAD 2.12 chain 6 (SEQ ID NO: 16) (MYH6) Myosin heavy Homo sapiens NP_002462.2 1939 SLKLMATLFSSYATAD 8.39 87.50 chain 6 (SEQ ID NO: 17) (MYH6) β- Bacteroides WP_109116112.1 1022 FLILMAALTATFASAQ 2.71 56.25 galactosidase faecis 1 (SEQ ID NO: 18) β- Bacteroides SQA30530.1 1023 FLILMAALTATFASAQ 2.71 56.25 galactosidase thetaiotaomicron 1 (SEQ ID NO: 19) Cysteine Enterobacter KJX05885.1 238 LIGMMSTESTYASAQE 16.78 50.00 hydrolase cloacae 1 (SEQ ID NO: 20) MFS Enterococcus AWM66597.1 475 GMTLMGLSTLFLSTYA 14.00 37.50 transporter faecalis 1 (SEQ ID NO: 21) Outer Chlamydia CAA39396.1 547 LETSMAEFTSTNVISL 21.87 25.00 membrane pneumoniae (SEQ ID NO: 22) protein 21Commensal in human intestina microbiome. 2Amino acids matching to murine MYH6614-628 are highlighted by underlining. 3MHC class II Iad binding prediction algorithm: IEDB analysis resource (low percentile rank indicates better binding to MHC II molecule). - The predicted binding of human MYH6614-629 and Bacteroides β-gal11-25 peptides to HLA-DQ alleles was performed using the full HLA reference set, which provides >99% of coverage of the HLA allele usage (40). Input sequences from full human myosin heavy chain alpha (myosin
heavy chain 6, MYH6) protein (accession number: NP_002462.2) and B. thetaiotaomicron β-galactosidase (accession number: SQA30530.1) were used for the prediction in IEDB. - The HLA-genotyping for the AMITIS patients was performed by the Institut für Klinische Transfusionsmedizin and Hämotherapie from the Universitätsklinikum Wurzburg. Briefly, genomic DNA was obtained from PBMCs and analyzed by a sequence based typing (SBT) by Sanger sequencing using forward and reverse primers for sequencing the
exons 2 and 3 (HLA class H) in a Applied Biosystems Genetic analyzer 3130×I, instrument (ThermoFisher Scientific), using SBTexcellerator kits (Gendx) and the analysis software SBTengine. The Micro-DCM cohort HLA typing was performed by Illumina sequencing (Histogenetics USA). - To detect anti-β1 antibodies in the sera of patients we used Dynabeads® M-270 Epoxy (Life Technologies) coated for 72 h according to the manufacturer's instructions with 100, 80, 60, 40, 20, 10, 5 or 2.5 μg/ml β1-ECII peptide. Control beads were coated with 10 μg/ml of a scramble peptide comprising the same amino acids of the β1-ECII peptide. After the coating the beads were washed using phosphate-buffered saline (PBS)/0.1% Bovine Serum Albumin (BSA) and stored in PBS/0.1% BSA/0.02% NaN3 until further use. For
antibody detection 104 beads of every fraction were seeded into a 96-well-V-bottom plate. After blocking the beads with phosphate-buffered saline (PBS)/10% Bovine Serum Albumin (BSA) (15 min, on ice), a 1:100 dilution in PBS/0.1% BSA/0.02% NaN3 of patient serum, or as negative control pooled human AB serum (Sigma), was incubated with the beads in a total volume of 50 μl (15 min, on ice). As a positive control the rat anti-β1-ECII monoclonal antibody 13F6 was used (10 μg/ml in PBS/0.1% BSA/0.02% NaN3/AB serum 1:100). The beads were washed four times using PBS/0.1% BSA/0.02% NaN3 and transferred to fresh wells before incubation with a 1:100 dilution of the secondary antibody F(ab)2 donkey-anti-human IgG (H+L) RTC (Dianova) in PBS/0.1% BSA/0.02% NaN3 (total volume: 50 μl, 15 min, on ice). 13F6 was detected using F(ab)2 donkey-anti-rat IgG (H+L) PE (Dianova; 1:1000 dilution in PBS/0.1% BSA/0.02% NaN3). After a final washing step the samples were measured on a FACScan flow cytometer and the data analyzed using FlowJo (Treestar). Half maximal binding was determined using GraphPad Prism. When a half maximal binding value was calculable, the serum was defined as antibody-positive. - IFN-γ ELISPOT was performed as described by Lv et. al. with minor modifications (7). Briefly, PBMCs from patients and healthy volunteers were isolated on density gradients. PBMCs were adjusted to 2×106 cells/ml in 0.5 ml of complete IVS medium supplemented with 8% of human serum. Cells were stimulated for 48 h with 0.25 μg/ml of the indicated peptide or left untreated as medium control. After incubation, cells were washed and re-suspended in 0.3 ml of RPMI/10% FCS and approximately 5×105 cells/0.1 ml were dispensed in duplicates into previously coated and blocked ELISPOT plates (IFN-γ base kit, MABTECH). After 24 h of incubation at 37° C. the ELISPOT plates were washed and developed as indicated by the manufacturer. Plates were counted using an ELISPOT reader and analyzed with the software ELISPOT 3.1SR (AID). Number of specific IFN-γ positive cells was calculated as the mean of the duplicates minus the mean of the medium control.
- Fecal microbiota transplantation was performed as previously reported by Gopalakrishnan et. al (41). Briefly, germ free TCRM and transgene-negative littermates received FMT from myocarditis patients which were previously confirmed to be positive for B. thetaiotaomicron and the DQB1*03:02 HLA allele; each patient sample was administered to 4 mice. 200 μl cleared supernatant from 0.1 g/μl human fecal suspension was obtained using a 100 μm strainer and gavaged into
mice 3 times over 1 week. B. thetaiotaomicron colonization was assessed by qPCR at 14 and 21 days after the first administration. - Statistical analyses were performed with Graphpad Prism 7.0 using unpaired two-tailed Student's t-test or Mann-Whitney U tests. Longitudinal comparison between different groups was performed by one-way ANOVA with Tukey's post-test or two-way ANOVA with Bonferroni's post-test. For allele comparison, the Fisher's exact test was used. Statistical significance was defined as p<0.05.
- The link between infection with pathogens and the development of autoimmune diseases is well-documented (17, 18), while the impact of commensal bacteria on dysregulated self-reactivity has only recently been uncovered (19-23). Importantly, commensal bacteria such as Bacteroides can also dampen autoimmune reactivity both in mice and humans (24, 25). To assess whether the intestinal environment and the vast repertoire of antigens present in the microbiota shape heart-specific autoimmunity, we used transgenic mice that express a MYH6-specific T cell receptor on more than 95% of their CD4+ T cells (TCRM) (8). As described previously (8), all TCRM mice developed spontaneous autoimmune myocarditis and approximately 50% of the animals progressed from autoimmune myocarditis to lethal DCM under specific pathogen free (SPF) housing conditions (
FIG. 1A-D ). In stark contrast, the lack of a commensal microbiome under germ-free (GF) conditions rescued TCRM mice from progression to the lethal disease (FIG. 1A ), abrogated cardiac dilatation (FIG. 1B ) and substantially reduced fibrotic remodeling of the cardiac tissue in 12 week old animals (FIG. 1C ). Histopathological assessment of disease severity revealed that myocarditis in GF TCRM mice was significantly reduced at 12 weeks when compared to age-matched TCRM mice raised under SPF conditions (FIG. 1D ). Echocardiographic analysis of heart functions with recording of the ejection fraction (FIG. 1E ), fractional shortening (FIG. 1F ) and systolic left ventricle internal diameter (LVID) (FIG. 1G ) revealed that hearts of 12 week old GF TCRM mice functioned normally despite the low level cardiac inflammation. The small difference in disease severity at 4 weeks of age and the lack of progressive myocarditis in GF TCRM mice (FIG. 1D ) suggested that post-weaning processes such as microbial colonization critically impact disease aggravation in TCRM mice. - To determine to which extent microbial colonization affected the activation of heart-specific CD4+ T cells, TCRM mice were transferred from GF to the SPF environment at 4 and 8 weeks of age and co-housed (CoH) with SPF TCRM mice (
FIG. 1H ). Colonization with the SPF microbiome precipitated progression to lethal disease in ex-GF mice within 6-12 weeks (FIG. 1I ), significantly exacerbated cardiac inflammation (FIG. 1J ) and impaired cardiac function (FIG. 5A-C ). Moreover, microbial colonization of GF TCRM mice led to rapid infiltration of the heart with CD45+ immune cells (FIG. 1K ) and an overshooting accumulation of MYH6-specific CD4+ T cells that expressed the transgenic Vβ8 chain (FIG. 1L ). While IFN-γ production of heart-infiltrating TCRM Th cells did not differ between SPF and GF mice (FIG. 1M ), the expression of IL-17 was dependent on the microbial status (FIG. 1N ). Co-housing increased the fraction of both IFN-γ-(FIG. 1M ) and IL-17-secreting heart-specific CD4+ T cells (FIG. 1N ) and promoted the accumulation of inflammatory myeloid cells in the heart tissue (FIG. 1O-R andFIG. 5D-F ) including inflammatory monocytes expressing CCR2 (FIG. 1P ), MHCIIhigh macrophages (FIG. 1Q ) and MERTK+ macrophages (FIG. 1R ). These data suggest that the presence of microbiota fosters the imprinting of a Th17 phenotype in heart-specific CD4+ T cells and favors the accumulation of inflammatory monocytes/macrophages in the myocardium of TCRM mice. - IL-17 determines the severity of experimental myocarditis in mice (6, 8) and appears to promote heart failure in human myocarditis patients (9). Since balancing of Th17 cell activity is crucial for the integrity and immune homeostasis of the intestinal lamina propria (26), we hypothesized that MYH6-specific CD4+ T cells in TCRM mice receive remote activation signals from the intestinal microbiota. Indeed, assessment of mucosal homing molecule expression on heart-infiltrating CD4+ TCRM cells revealed significantly elevated integrin α4β7 and chemokine receptor CCR6 expression under SPF compared to GF conditions (
FIG. 2A ). Adoptive transfer of dye-labeled TCRM cells into Rag1-deficient BALB/c mice (FIG. 6A ) revealed homing of CD4+ cells to the T cell zones of colonic patches (FIG. 2B ), colonic lymph nodes (FIG. 6B ) and mesenteric lymph nodes (FIG. 6C ). The adoptively transferred Vβ8+ CD4+ T cells showed detectable proliferation onday 3 in the lamina propria, the colonic lymph node and the mediastinal lymph node, while heart-infiltrating TCRM cells could only be detected on day 7 (FIG. 2C andFIG. 6D ). The CD4+ T cell response in the hearts of 4 week old TCRM mice was dominated by IFN-γ-producing Th cells (FIG. 6E ), whereas the colonic lamina propria environment fostered the differentiation of IL-17-producing CD4+ T cells (FIG. 6E and F). Importantly, only MYH6-specific CD4+ T cells, but not ovalbumin-specific CD4+ T cells from the colonic lamina propria were able to secrete IFN-γ or IL-17 upon ex-vivo re-stimulation with the MYH6 peptide (FIG. 6G ). These data indicate that heart-specific Th cells can be activated in the colonic lamina propria and/or the associated lymphoid organs prior to receiving further antigenic stimulation in the heart. - To further explore the interaction of heart-specific Th cells and the intestinal microbiome, 4 week old GF TCRM mice were co-housed with SPF transgene-negative (Tg−) and SPF TCRM littermates (
FIG. 2D ). Next generation sequencing of 16S rRNA genes amplified from feces revealed that co-housed Tg− and TCRM mice possess disparate microbiomes (FIG. 6H ) and that the microbiome of co-housed ex-GF TCRM mice appears to be more similar to the SPF TCRM microbiome (FIG. 2E ). More detailed analyses confirmed that GF TCRM mice acquired the microbiome of SPF TCRM mice (FIG. 2F ) with particularly high changes in the genera Bacteroides and Parabacteroides (FIG. 6I ). These data suggested that MYH6-specific CD4+ T cells in the TCRM mice impact the composition of the microbiome through antigen-specific interactions. Indeed, an in silico search for potentially cross-reactive epitopes identified two mimic peptides with high similarity to MYH6 in the β-galactosidase (β-gal) of Bacteroides thetaiotaomicron (B. theta) and B. faecis that possesses a homology of 56%, and cysteine hydrolase peptide of Enterobacter cloacae with 50% homology (Table 1). The B. theta β-gal peptide but not the Enterobacter cloacae peptide, induced in vitro proliferation of CD4+ T cells from TCRM mice (FIG. 2G ), while ovalbumin-specific CD4+ T cells were not activated by the microbial peptides (FIG. 7A ). TCRM CD4+ T cells exhibit a functional avidity of 6.3×10−6 M for the cross-reactive B. theta peptide (FIG. 7B ) that facilitated efficient activation of TCRM T cells in vivo by B. theta peptide-pulsed dendritic cells (FIG. 6C ) and ex vivo re-stimulation of transgenic CD4+ T cells from heart and colon (FIG. 2H ) of 12 week old TCRM mice. To elucidate the role of B. theta for the activation of TCRM Th cells in the colonic lamina propria, we mono-colonized GF TCRM mice with B. theta or Escherichia (E.). coli, or colonized them with our SPF microbiota (FIG. 7D ). These analyses showed that only B. theta mono-colonization specifically fostered the induction of IL-17 production as seen following SPF colonization (FIG. 7E ). The differentiation towards IFN-γ-producing CD4+ T cells was not significantly affected by any of the re-colonization conditions (FIG. 7E ). Next, we generated a B. theta mutant lacking the β-gal BT1626 gene encoding for the MYH6 peptide mimic (B. theta β-gal) (FIG. 7F ). Mono-colonization of GF TCRM mice with B. theta β-gal or the parental strain (FIG. 2I ) resulted in efficient seeding of the intestinal microenvironments with both strains (FIG. 7G ), but precipitated significantly reduced accumulation of CD45+ immune cells and Vβ8+CD4+ T cells in the myocardium (FIG. 2J ). Moreover, IL-17 production of myosin-specific CD4+ T cells in heart and colon was significantly reduced when TCRM mice were re-colonized with B. theta β-gal (FIG. 2K ). Assessment of IgA binding to intestinal bacteria using bacterial flow cytometry (FIG. 7H ) indicated that the presence of TCRM cells substantially increases the production of IgA against commensal microbiota under SPF conditions (FIG. 7I ). To determine to which extent such increased intestinal IgA production depends on specific antigenic stimulation by the MYH6 mimic peptide, we assessed B. theta-specific IgA binding in GF TCRM mice that were mono-colonized with B. theta β-gal. Indeed, the absence of the cross-reactive epitope in B. theta β-gal led to a significantly reduced IgA response when compared to re-colonization with the parental B. theta strain (FIG. 2L and M). In sum, these data reveal that organ-specific, genuinely autoreactive CD4+ T cells can shape the colonic microbiome and that distinct bacterial communities, here Bacteroides, represent a source of mimic peptides that can activate MYH6-specific CD4+ T cells. - Since the microbiome can be manipulated through the utilization of antibiotics, we assessed whether reduction of Bacteroides communities by broad-spectrum antibiotics combination (Sulfadoxine, Trimethoprim and Metronidazol; S+T+M) affects disease progression in TCRM mice. We found that antibiotics treatment resulted in a substantial shift in the microbiome composition including the reduced abundance of Bacteroides (
FIG. 8A ). None of the treated TCRM mice succumbed to lethal inflammatory cardiomyopathy (FIG. 3A ), which was most likely due to significantly reduced cardiac inflammation under antibiotic treatment (FIG. 3B and C). To corroborate these findings and to dissect the activation patterns of cardiac myosin-specific T cells, we utilized the adoptive transfer model of TCRM splenocytes into Rag1−/− mice with antibiotics treatment starting one week before the splenocyte transfer (FIG. 3D ). As expected, treatment with broad-spectrum antibiotics such as Vancomycin reduced not only B. theta levels in fecal samples (FIG. 3E ), but significantly ameliorated the cardiac disease atday 28 post adoptive transfer (FIG. 3F ). Flow cytometric analysis confirmed the profound reduction of the cardiac inflammation (FIG. 3G ) with significantly reduced accumulation of TCR-transgenic CD4+ T cells in hearts of Rag1−/− recipients treated with Streptomycin, S+T+M or Vancomycin (FIG. 3H ). Likewise, these three antibiotic regimens reduced CD45+ immune cell (FIG. 8B ) and Vβ8+ CD4+ T cell accumulation in the colon (FIG. 8C ), but not in the spleen of Rag1−/− recipients (FIG. 8D ). Vancomycin and S+T+M antibiotics treatment had a pronounced impact on the activation of heart-infiltrating and colonic Th17 cells, while the activity of Th1 cells was reduced to a lesser extent (FIG. 3I andFIG. 8E-F ). The adoptively transferred B cells from TCRM spleens generated B. theta-specific IgG antibodies that could be detected in the serum of Rag1−/− recipients onday 28 post transfer (FIG. 3J ). Antibiotics treatment reduced B. theta-specific IgG antibodies, but had variable effects on IgG antibody responses against B. distasonis, B. vulgatus or Enterobacter cloacae (FIG. 3J ) indicating that antibiotics treatment did not negatively impact global immune reactivity in the Rag1−/− recipients. These results and the finding that adoptive transfer of heart-specific TCRM cells to BALB/c mice significantly enhanced anti-B. theta antibody responses (FIG. 3K ) further support the notion that heart-specific CD4+ T cells specifically interact with microbial components in the intestine and thereby impact systemic immune reactivity. - To establish translational relevance of the link between microbial molecular mimicry and inflammatory cardiomyopathy, we assessed anti-Bacteroides IgG responses in sera from patients with endomyocardial biopsy-proven myocarditis (Etics/AMITIS study, Table 2) (27). Patients with acute myocarditis showed significantly elevated B. theta-specific IgG responses compared to a healthy control group (
FIG. 4A ). Clinical improvement of the myocarditis patients with gain of heart function (FIG. 4B ) and reduced inflammatory status (FIG. 4C ) was accompanied by a reduction in seroreactivity against B. theta (FIG. 4A ). Based on anti-B. theta IgG reactivity in patients with acute myocarditis, we grouped the patients into high (>mean, red box inFIG. 4A ) and low (<mean, black box inFIG. 4A ) responders and correlated their antibody status with the available clinical parameters: positivity for anti-beta 1 adrenergic receptor autoantibodies (FIG. 9A ), C-reactive protein (CRP) levels (FIG. 9B ) and an ejection fraction cut-off of 40% (FIG. 9C ). Acute myocarditis patients with low anti-B. theta reactivity showed a combined clinical score that was significantly lower compared to the high responder group (FIG. 4D ) indicating that systemic anti-Bacteroides immune reactivity is linked to myocardial inflammation in humans. - While IgG antibodies against B. theta in the AMITIS cohort were diminished at subsequent visits (
FIG. 4A ), anti-Enterobacter cloacae (FIG. 9D ), anti-Escherichia coli antibodies (FIG. 9E ) and total serum IgG concentrations (FIG. 9F ) did not change significantly. Moreover, IgG antibody reactivity against other Bacteroides species (B. distasonis and B. vulgatus) was not different in AMITIS myocarditis patients and healthy controls (FIG. 4E ). To corroborate these results, we established a prospective clinical study and included thirteen acute myocarditis patients showing typical late enhancement pattern in cardiac magnetic resonance imaging and four acute heart failure patients with left ventricular ejection fraction ≤40% (Micro-DCM study; Table 3). A cohort of age-matched healthy volunteers served as controls (Table 4). Importantly, myocarditis patients of the Micro-DCM study showed significantly elevated anti-B. theta IgG antibodies (FIG. 4F ), while seroreactivity against other bacteria—except reduced reactivity against B. vulgatus—were not different when compared to healthy controls (FIG. 4G ). - Preclinical studies in mice have established that MYH6 is the dominant autoantigen in the heart tissue (5, 28) and that susceptibility to experimental autoimmune myocarditis depends mainly on the MHC haplotype, whereby MYH6 epitopes appear to be presented by different MHC class II molecules (29). Bioinformatics analysis of binding affinities of the B. theta β-gal11-25 peptide mimic indicated several HLA-DQA1*/HLA-DQB1* combinations, including HLA-DQ8, that are also predicted to bind human MYH6614-629 (
FIG. 4H ). Assessment of T cell reactivity against the MYH6 and β-gal peptides in peripheral blood mononuclear cells Micro-DCM patients revealed that patients exhibit significantly higher IFN- reactivity against both peptides when compared to healthy controls (FIG. 4I ). Moreover, the frequency of IFN--producing T cells in Micro-DCM patients that possess alleles other than HLA-DQB1* 03 was also reduced in comparison to the HLA-DQB1*03-positive group (FIG. 4I ). When we focused the analysis on Micro-DCM myocarditis patients and individuals showing a T cell response >0 in the ELISpot assay for at least one of the peptides, we found a highly significant correlation between MYH6 and β-gal11-25 peptide reactivity (FIG. 9K ) suggesting that heart-specific CD4+ T cells cross-react with the bacterial peptide in human myocarditis patients. Finally, to provide evidence that the presence of B. theta in the human intestinal microbiome is causally linked to myocarditis development, we performed fecal transplants from B. theta-positive myocarditis patients into GF TCRM and transgene-negative mice (FIG. 4J ). Both groups of mice were equally well colonized by B. theta (FIG. 4K ). Increased accumulation of CD45+ immune cells (FIG. 4L ) and CD4+ T cells (FIG. 4M ) in TCRM hearts after four weeks of re-colonization indicates that the microbiome of human myocarditis patients contains the pathological principle that drives myocardial inflammation in TCRM mice. In sum, the causal evidence provided by the preclinical studies in TCRM mice and the significant correlation of systemic anti-Bacteroides immune reactivity with disease severity in human myocarditis patients indicate that inflammatory cardiomyopathy in humans is driven—at least partially—through the activation of heart-specific Th cells by bacterial peptide mimics that are derived from the intestinal microbiota. -
- 1. J. Buggey, C. A. ElAmm, Myocarditis and cardiomyopathy. Curr Opin Cardiol 33, 341-346 (2018).
- 2. R. G. Weintraub, C. Semsarian, P. Macdonald, Dilated cardiomyopathy. Lancet 390, 400-414 (2017).
- 3. N. R. Rose, Learning from myocarditis: mimicry, chaos and black holes.
F 1000Prime Rep 6, 25 (2014). - 4. B. H. Trachtenberg, J. M. Hare, Inflammatory Cardiomyopathic Syndromes. Circ Res 121, 803-818 (2017).
- 5. P. Krebs et al., Molecular mapping of autoimmune B cell responses in experimental myocarditis.
J. Autoimmun 28, 224-233 (2007). - 6. M. Rangachari et al., T-bet negatively regulates autoimmune myocarditis by suppressing local production of
interleukin 17. J. Exp. Med 203, 2009-2019 (2006). - 7. H. Lv et al., Impaired thymic tolerance to alpha-myosin directs autoimmunity to the heart in mice and humans. J. Clin. Invest 121, 1561-1573 (2011).
- 8. V. Nindl et al., Cooperation of Th1 and Th17 cells determines transition from autoimmune myocarditis to dilated cardiomyopathy.
Eur J Immunol 42, 2311-2321 (2012). - 9. J. M. Myers et al., Cardiac myosin-Th17 responses promote heart failure in human myocarditis.
JCI Insight 1, (2016). - 10. B. Maisch, P. Alter, Treatment options in myocarditis and inflammatory cardiomyopathy: Focus on i.v. immunoglobulins. Herz, (2018).
- 11. S. Heymans, U. Eriksson, J. Lehtonen, L. T. Cooper, Jr., The Quest for New Approaches in Myocarditis and Inflammatory Cardiomyopathy. J Am Coll Cardiol 68, 2348-2364 (2016).
- 12. E. Generali, A. Ceribelli, M. A. Stazi, C. Selmi, Lessons learned from twins in autoimmune and chronic inflammatory diseases. J Autoimmun 83, 51-61 (2017).
- 13. A. Davidson, B. Diamond, Autoimmune diseases. N Engl J Med 345, 340-350 (2001).
- 14. W. Liu, W. M. Li, N. L. Sun, HLA-DQA1, -DQB1 polymorphism and genetic susceptibility to idiopathic dilated cardiomyopathy in Hans of northern China. Ann Hum Genet 69, 382-388 (2005).
- 15. I. Portig, A. Sandmoeller, S. Kreilinger, B. Maisch, HLA-DQB1* polymorphism and associations with dilated cardiomyopathy, inflammatory dilated cardiomyopathy and myocarditis.
Autoimmunity 42, 33-40 (2009). - 16. J. A. Taylor et al., A spontaneous model for autoimmune myocarditis using the human MHC molecule HLA-DQ8. J. Immunol 172, 2651-2658 (2004).
- 17. C. L. Vanderlugt, S. D. Miller, Epitope spreading in immune-mediated diseases: implications for immunotherapy.
Nat. Rev. Immunol 2, 85-95 (2002). - 18. M. B. Oldstone, Molecular mimicry and autoimmune disease.
Cell 50, 819-820 (1987). - 19. F. Teng et al., Gut Microbiota Drive Autoimmune Arthritis by Promoting Differentiation and Migration of Peyer's Patch T Follicular Helper Cells. Immunity 44, 875-888 (2016).
- 20. R. Horai et al., Microbiota-Dependent Activation of an Autoreactive T Cell Receptor Provokes Autoimmunity in an Immunologically Privileged Site. Immunity 43, 343-353 (2015).
- 21. Y. K. Lee, J. S. Menezes, Y. Umesaki, S. K. Mazmanian, Proinflammatory T-cell responses to gut microbiota promote experimental autoimmune encephalomyelitis. Proc Natl
Acad Sci USA 108Suppl 1, 4615-4622 (2011). - 22. Y. E. Chen, M. A. Fischbach, Y. Belkaid, Skin microbiota-host interactions. Nature 553, 427-436 (2018).
- 23. N. Tai et al., Microbial antigen mimics activate diabetogenic CD8 T cells in NOD mice. J Exp Med 213, 2129-2146 (2016).
- 24. T. Vatanen et al., Variation in Microbiome LPS Immunogenicity Contributes to Autoimmunity in Humans. Cell 165, 842-853 (2016).
- 25. R. Hebbandi Nanjundappa et al., A Gut Microbial Mimic that Hijacks Diabetogenic Autoreactivity to Suppress Colitis. Cell 171, 655-667 e617 (2017).
- 26. K. Honda, D. R. Littman, The microbiota in adaptive immune homeostasis and disease. Nature 535, 75-84 (2016).
- 27. N. Deubner et al., Cardiac beta1-adrenoceptor autoantibodies in human heart disease: rationale and design of the Etiology, Titre-Course, and Survival (ETiCS) Study. Eur J Heart Fail 12, 753-762 (2010).
- 28. C. L. Pummerer et al., Identification of cardiac myosin peptides capable of inducing autoimmune myocarditis in BALB/c mice. J. Clin. Invest 97, 2057-2062 (1996).
- 29. J. G. Barin, D. Cihakova, Control of inflammatory heart disease by CD4+ T cells. Ann N Y Acad Sci 1285, 80-96 (2013).
- 30. L. Zhang et al., Cardiotoxicity of Immune Checkpoint Inhibitors. Curr Treat
Options Cardiovasc Med 21, 32 (2019). - 31. D. B. Johnson et al., Fulminant Myocarditis with Combination Immune Checkpoint Blockade. N Engl J Med 375, 1749-1755 (2016).
- 32. A. Cossarizza et al., Guidelines for the use of flow cytometry and cell sorting in immunological studies. Eur J Immunol 47, 1584-1797 (2017).
- 33. N. M. Koropatkin, E. C. Martens, J. I. Gordon, T. J. Smith, Starch catabolism by a prominent human gut symbiont is directed by the recognition of amylose helices.
Structure 16, 1105-1115 (2008). - 34. M. Mamantopoulos et al., NIrp6- and ASC-Dependent Inflammasomes Do Not Shape the Commensal Gut Microbiota Composition. Immunity 47, 339-348 e334 (2017).
- 35. T. Miki, R. Goto, M. Fujimoto, N. Okada, W. D. Hardt, The Bactericidal Lectin Reglllbeta Prolongs Gut Colonization and Enteropathy in the Streptomycin Mouse Model for Salmonella Diarrhea.
Cell Host Microbe 21, 195-207 (2017). - 36. J. J. Bunker et al., Innate and Adaptive Humoral Responses Coat Distinct Commensal Bacteria with Immunoglobulin A. Immunity 43, 541-553 (2015).
- 37. C. Gil-Cruz et al., Fibroblastic reticular cells regulate intestinal inflammation via IL-15-mediated control of
group 1 ILCs.Nat Immunol 17, 1388-1396 (2016). - 38. J. G. Caporaso et al., QIIME allows analysis of high-throughput community sequencing data.
Nat Methods 7, 335-336 (2010). - 39. C. A. Hickey et al., Colitogenic Bacteroides thetaiotaomicron Antigens Access Host Immune Cells in a Sulfatase-Dependent Manner via Outer Membrane Vesicles.
Cell Host Microbe 17, 672-680 (2015). - 40. J. Greenbaum et al., Functional classification of class II human leukocyte antigen (HLA) molecules reveals seven different supertypes and a surprising degree of repertoire sharing across supertypes. Immunogenetics 63, 325-335 (2011).
- 41. V. Gopalakrishnan et al., Gut microbiome modulates response to anti-PD-1 immunotherapy in melanoma patients. Science 359, 97-103 (2018).
Claims (20)
1. A method of preventing myocarditis in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
2. A method of treating myocarditis or DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
3. A method of limiting progression of myocarditis toward DCM in a subject in need thereof, comprising reducing the amount of Bacteroides sp. in a subject.
4. A method of diagnosis a subject as having myocarditis or DCM, comprising obtaining a biological sample of the subject, quantifying the amount of Bacteroides sp. in the biological sample relative to a control sample.
5. The method of claim 1 , comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
6. The method of claim 2 , comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
7. The method of claim 3 , comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
8. The method of claim 4 , comprising administering to the subject an effective amount of an antibiotic, phage, recombinant phage, packaged phagemid, bacteria, engineered bacteria, bacteriocin or endolysin.
9. The method of claim 5 , wherein the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
10. The method of claim 6 , wherein the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
11. The method of claim 7 , wherein the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
12. The method of claim 8 , wherein the antibiotic is selected from streptomycin, vancomycin, clindamycin, metronidazole, sulphadoxine, trimethoprim, or any combination of 1, 2, 3, 4, 5 or 6 of these antibiotics.
13. The method of claim 5 , wherein the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
14. The method of claim 6 , wherein the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
15. The method of claim 7 , wherein the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
16. The method of claim 8 , wherein the phage, recombinant phage or packaged phagemid encodes a nuclease selected from CRISPR-Cas, TALENs and variants, zinc finger nuclease (ZFN) and ZFN variants, natural, evolved or engineered meganuclease or recombinase variants.
17. The method of claim 5 , wherein the Bacteroides is B. thetaiotaomicron and/or B. faecis.
18. The method of claim 6 , wherein the Bacteroides is B. thetaiotaomicron and/or B. faecis.
19. The method of claim 7 , wherein the Bacteroides is B. thetaiotaomicron and/or B. faecis.
20. The method of claim 8 , wherein the Bacteroides is B. thetaiotaomicron and/or B. faecis.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US18/159,482 US20230226129A1 (en) | 2020-04-08 | 2023-01-25 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US202063007197P | 2020-04-08 | 2020-04-08 | |
US17/225,854 US11617773B2 (en) | 2020-04-08 | 2021-04-08 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
US18/159,482 US20230226129A1 (en) | 2020-04-08 | 2023-01-25 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
Related Parent Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
US17/225,854 Division US11617773B2 (en) | 2020-04-08 | 2021-04-08 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
Publications (1)
Publication Number | Publication Date |
---|---|
US20230226129A1 true US20230226129A1 (en) | 2023-07-20 |
Family
ID=78006404
Family Applications (2)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
US17/225,854 Active US11617773B2 (en) | 2020-04-08 | 2021-04-08 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
US18/159,482 Abandoned US20230226129A1 (en) | 2020-04-08 | 2023-01-25 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
Family Applications Before (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
US17/225,854 Active US11617773B2 (en) | 2020-04-08 | 2021-04-08 | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy |
Country Status (1)
Country | Link |
---|---|
US (2) | US11617773B2 (en) |
Family Cites Families (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5863560A (en) | 1996-09-11 | 1999-01-26 | Virotex Corporation | Compositions and methods for topical application of therapeutic agents |
GB0209680D0 (en) | 2002-04-27 | 2002-06-05 | Univ Strathclyde | Immobilisation and stabilisation of bacteriophage |
US20110218216A1 (en) | 2010-01-29 | 2011-09-08 | Kumaravel Vivek | Extended release pharmaceutical composition of donepezil |
US20150166982A1 (en) | 2013-12-12 | 2015-06-18 | President And Fellows Of Harvard College | Methods for correcting pi3k point mutations |
AU2017306676B2 (en) | 2016-08-03 | 2024-02-22 | President And Fellows Of Harvard College | Adenosine nucleobase editors and uses thereof |
WO2020181193A1 (en) | 2019-03-06 | 2020-09-10 | The Broad Institute, Inc. | T:a to a:t base editing through adenosine methylation |
WO2020181180A1 (en) | 2019-03-06 | 2020-09-10 | The Broad Institute, Inc. | A:t to c:g base editors and uses thereof |
WO2020181178A1 (en) | 2019-03-06 | 2020-09-10 | The Broad Institute, Inc. | T:a to a:t base editing through thymine alkylation |
WO2020181202A1 (en) | 2019-03-06 | 2020-09-10 | The Broad Institute, Inc. | A:t to t:a base editing through adenine deamination and oxidation |
WO2020181195A1 (en) | 2019-03-06 | 2020-09-10 | The Broad Institute, Inc. | T:a to a:t base editing through adenine excision |
-
2021
- 2021-04-08 US US17/225,854 patent/US11617773B2/en active Active
-
2023
- 2023-01-25 US US18/159,482 patent/US20230226129A1/en not_active Abandoned
Also Published As
Publication number | Publication date |
---|---|
US20210315950A1 (en) | 2021-10-14 |
US11617773B2 (en) | 2023-04-04 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Price et al. | A map of toll-like receptor expression in the intestinal epithelium reveals distinct spatial, cell type-specific, and temporal patterns | |
US20230330157A1 (en) | Modulation of microbiota function by gene therapy of the microbiome to prevent, treat or cure microbiome-associated diseases or disorders | |
JP2023101670A (en) | Intestinal microbiota and gvhd | |
Bromberg et al. | Gut microbiota–dependent modulation of innate immunity and lymph node remodeling affects cardiac allograft outcomes | |
Nieuwenhuis et al. | Cd1d-dependent regulation of bacterial colonization in the intestine of mice | |
Schumacher et al. | Gastric Sonic Hedgehog acts as a macrophage chemoattractant during the immune response to Helicobacter pylori | |
JP2022046822A (en) | Compositions and methods for treating diseases and disorders of central nervous system | |
Lochhead et al. | Robust interferon signature and suppressed tissue repair gene expression in synovial tissue from patients with postinfectious, Borrelia burgdorferi‐induced Lyme arthritis | |
US11712436B2 (en) | Theranostic test for antifungal treatment of inflammatory diseases | |
Malachowa et al. | Vaccine protection against multidrug-resistant Klebsiella pneumoniae in a nonhuman primate model of severe lower respiratory tract infection | |
US20220112280A1 (en) | Transplant tolerance induction with carbodiimide treated tolerizing vaccine | |
JP2007523640A (en) | Methods for inducing or modulating an immune response | |
JP6346602B2 (en) | Isolation and use of inhibitory stromal cells from human lymphoid organs | |
Liu et al. | Histone deacetylase 9 deficiency exaggerates uterine M2 macrophage polarization | |
Kohl et al. | Macrophages inhibit Coxiella burnetii by the ACOD1‐itaconate pathway for containment of Q fever | |
Jin et al. | Antigen-presenting aged neutrophils induce CD4+ T cells to exacerbate inflammation in sepsis | |
Snell et al. | Consequences of donor-derived passengers (pathogens, cells, biological molecules and proteins) on clinical outcomes | |
US11617773B2 (en) | Elimination of colonic bacterial driving lethal inflammatory cardiomyopathy | |
Ebrahimnezhaddarzi et al. | Mpeg1 is not essential for antibacterial or antiviral immunity, but is implicated in antigen presentation | |
US20230201323A1 (en) | Vaccine formulations | |
US20190374632A1 (en) | Compositions and methods of treating autoimmune disease by reducing enterococcus | |
JP2001526043A (en) | TGC-methods for inducing targeted somatic cell transgenesis | |
Amato-Menker et al. | XX sex chromosome complement modulates immune responses to heat-killed Streptococcus pneumoniae immunization in a microbiome-dependent manner | |
US20170232092A1 (en) | Methods and compositions for t cell generation and uses thereof | |
JP7411970B2 (en) | Agent for increasing IgA antibody content in milk |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
STCB | Information on status: application discontinuation |
Free format text: ABANDONED -- INCOMPLETE APPLICATION (PRE-EXAMINATION) |