NZ620075B2 - A method for predicting clinical benefit in the treatment of neurodevelopmental, neurological or neuropsychiatric disorders - Google Patents
A method for predicting clinical benefit in the treatment of neurodevelopmental, neurological or neuropsychiatric disorders Download PDFInfo
- Publication number
- NZ620075B2 NZ620075B2 NZ620075A NZ62007512A NZ620075B2 NZ 620075 B2 NZ620075 B2 NZ 620075B2 NZ 620075 A NZ620075 A NZ 620075A NZ 62007512 A NZ62007512 A NZ 62007512A NZ 620075 B2 NZ620075 B2 NZ 620075B2
- Authority
- NZ
- New Zealand
- Prior art keywords
- variant
- cfhr1
- protein
- complement factor
- patients
- Prior art date
Links
- 201000010099 disease Diseases 0.000 title abstract description 54
- 230000001123 neurodevelopmental Effects 0.000 title abstract description 36
- 230000000926 neurological Effects 0.000 title abstract description 34
- 108010053085 Complement Factor H Proteins 0.000 claims abstract description 214
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 139
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 139
- 230000004044 response Effects 0.000 claims abstract description 63
- DHMQDGOQFOQNFH-UHFFFAOYSA-N glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 claims abstract description 46
- 238000000338 in vitro Methods 0.000 claims abstract description 37
- 102000016550 Complement Factor H Human genes 0.000 claims abstract description 30
- 239000004471 Glycine Substances 0.000 claims abstract description 23
- 230000002068 genetic Effects 0.000 claims abstract description 21
- 230000002401 inhibitory effect Effects 0.000 claims abstract description 21
- 239000003112 inhibitor Substances 0.000 claims abstract description 19
- 102100016300 CFH Human genes 0.000 claims description 186
- 239000000523 sample Substances 0.000 claims description 74
- 238000004166 bioassay Methods 0.000 claims description 48
- 201000000980 schizophrenia Diseases 0.000 claims description 43
- 102100015633 CFHR1 Human genes 0.000 claims description 39
- 101710002639 CFHR1 Proteins 0.000 claims description 39
- 108090001123 antibodies Proteins 0.000 claims description 36
- 102000004965 antibodies Human genes 0.000 claims description 36
- 238000005259 measurement Methods 0.000 claims description 28
- 239000003550 marker Substances 0.000 claims description 23
- 239000000902 placebo Substances 0.000 claims description 14
- 229940068196 placebo Drugs 0.000 claims description 14
- 230000027455 binding Effects 0.000 claims description 11
- 150000002500 ions Chemical class 0.000 claims description 9
- 238000002965 ELISA Methods 0.000 claims description 8
- 102100015632 CFHR3 Human genes 0.000 claims description 7
- 101710002647 CFHR3 Proteins 0.000 claims description 7
- -1 [4-(3-fluorotrifluormethyl-pyridinyl)-piperazinyl]-[5- methanesulfonyl[[(2S)-1,1,1-trifluoropropanyl]oxy]phenyl]methanone Chemical group 0.000 claims description 7
- 238000005516 engineering process Methods 0.000 claims description 3
- 201000000978 schizoaffective disease Diseases 0.000 claims description 2
- 108010071253 factor H-related protein 1 Proteins 0.000 claims 2
- SVPXDRXYRYOSEX-UHFFFAOYSA-N Bentoquatam Chemical compound O.O=[Si]=O.O=[Al]O[Al]=O SVPXDRXYRYOSEX-UHFFFAOYSA-N 0.000 claims 1
- 101700034586 CNMD Proteins 0.000 claims 1
- 239000000203 mixture Substances 0.000 abstract description 31
- 230000037361 pathway Effects 0.000 abstract description 22
- 230000000848 glutamatergic Effects 0.000 abstract description 20
- 150000001875 compounds Chemical class 0.000 abstract description 19
- 210000002966 Serum Anatomy 0.000 description 49
- 238000004458 analytical method Methods 0.000 description 26
- 102000004868 N-methyl-D-aspartate receptors Human genes 0.000 description 16
- 108090001041 N-methyl-D-aspartate receptors Proteins 0.000 description 16
- 229920003013 deoxyribonucleic acid Polymers 0.000 description 15
- 238000003018 immunoassay Methods 0.000 description 15
- 239000012071 phase Substances 0.000 description 15
- FAPWRFPIFSIZLT-UHFFFAOYSA-M sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 14
- 102100015637 CFHR5 Human genes 0.000 description 13
- 101710002642 CFHR5 Proteins 0.000 description 13
- 210000002381 Plasma Anatomy 0.000 description 13
- 239000011230 binding agent Substances 0.000 description 13
- 239000000090 biomarker Substances 0.000 description 13
- 210000004369 Blood Anatomy 0.000 description 12
- 102100015634 CFHR2 Human genes 0.000 description 12
- 101710002637 CFHR2 Proteins 0.000 description 12
- 239000008280 blood Substances 0.000 description 12
- 238000000034 method Methods 0.000 description 12
- 230000037165 Serum Concentration Effects 0.000 description 10
- 230000000694 effects Effects 0.000 description 10
- 201000008430 obsessive-compulsive disease Diseases 0.000 description 10
- 210000004027 cells Anatomy 0.000 description 9
- 239000011859 microparticle Substances 0.000 description 9
- 210000001124 Body Fluids Anatomy 0.000 description 8
- 210000004556 Brain Anatomy 0.000 description 8
- 239000010839 body fluid Substances 0.000 description 8
- 230000014509 gene expression Effects 0.000 description 8
- 108010045030 monoclonal antibodies Proteins 0.000 description 8
- 102000005614 monoclonal antibodies Human genes 0.000 description 8
- 239000011780 sodium chloride Substances 0.000 description 8
- 206010003805 Autism Diseases 0.000 description 7
- 241000282326 Felis catus Species 0.000 description 7
- 201000002055 autistic disease Diseases 0.000 description 7
- 230000000295 complement Effects 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 239000012491 analyte Substances 0.000 description 6
- 230000002596 correlated Effects 0.000 description 6
- 229940079593 drugs Drugs 0.000 description 6
- 239000007787 solid Substances 0.000 description 6
- 229920000272 Oligonucleotide Polymers 0.000 description 5
- 238000004164 analytical calibration Methods 0.000 description 5
- 238000011088 calibration curve Methods 0.000 description 5
- 238000001514 detection method Methods 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 230000018109 developmental process Effects 0.000 description 5
- 239000003085 diluting agent Substances 0.000 description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 5
- 238000000746 purification Methods 0.000 description 5
- 229940049906 Glutamate Drugs 0.000 description 4
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 4
- 102100014158 SLC6A9 Human genes 0.000 description 4
- 108060007768 SLC6A9 Proteins 0.000 description 4
- 230000005540 biological transmission Effects 0.000 description 4
- ZPUCINDJVBIVPJ-LJISPDSOSA-N cocaine Chemical compound O([C@H]1C[C@@H]2CC[C@@H](N2C)[C@H]1C(=O)OC)C(=O)C1=CC=CC=C1 ZPUCINDJVBIVPJ-LJISPDSOSA-N 0.000 description 4
- 229960003920 cocaine Drugs 0.000 description 4
- ZPUCINDJVBIVPJ-BARDWOONSA-N cocaine Natural products O([C@@H]1C[C@H]2CC[C@H](N2C)[C@@H]1C(=O)OC)C(=O)C1=CC=CC=C1 ZPUCINDJVBIVPJ-BARDWOONSA-N 0.000 description 4
- 230000000875 corresponding Effects 0.000 description 4
- 230000001419 dependent Effects 0.000 description 4
- 238000010790 dilution Methods 0.000 description 4
- 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 4
- 230000001404 mediated Effects 0.000 description 4
- 102000005962 receptors Human genes 0.000 description 4
- 108020003175 receptors Proteins 0.000 description 4
- 230000035945 sensitivity Effects 0.000 description 4
- 201000006152 substance dependence Diseases 0.000 description 4
- 206010004938 Bipolar disease Diseases 0.000 description 3
- 102100015636 CFHR4 Human genes 0.000 description 3
- 101710002644 CFHR4 Proteins 0.000 description 3
- 108009000280 Complement Activation Proteins 0.000 description 3
- 230000035693 Fab Effects 0.000 description 3
- 239000012505 Superdex™ Substances 0.000 description 3
- 229960000070 antineoplastic Monoclonal antibodies Drugs 0.000 description 3
- 230000006399 behavior Effects 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 230000024203 complement activation Effects 0.000 description 3
- 238000003745 diagnosis Methods 0.000 description 3
- 238000011156 evaluation Methods 0.000 description 3
- 230000036541 health Effects 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 229960000060 monoclonal antibodies Drugs 0.000 description 3
- 239000002858 neurotransmitter agent Substances 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 238000003753 real-time PCR Methods 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 206010064930 Age-related macular degeneration Diseases 0.000 description 2
- 206010002942 Apathy Diseases 0.000 description 2
- 229940098773 Bovine Serum Albumin Drugs 0.000 description 2
- 108091003117 Bovine Serum Albumin Proteins 0.000 description 2
- 102100012036 CCL23 Human genes 0.000 description 2
- 101700065817 CCL23 Proteins 0.000 description 2
- 238000007399 DNA isolation Methods 0.000 description 2
- 229940088598 Enzyme Drugs 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 206010018370 Glomerulonephritis membranoproliferative Diseases 0.000 description 2
- 102000003886 Glycoproteins Human genes 0.000 description 2
- 108090000288 Glycoproteins Proteins 0.000 description 2
- 101710015954 HVA1 Proteins 0.000 description 2
- 241000282619 Hylobates lar Species 0.000 description 2
- 108010054477 Immunoglobulin Fab Fragments Proteins 0.000 description 2
- 102000001706 Immunoglobulin Fab Fragments Human genes 0.000 description 2
- 206010022114 Injury Diseases 0.000 description 2
- 101700065814 LEA2 Proteins 0.000 description 2
- 101700021338 LEC Proteins 0.000 description 2
- 101700077545 LECC Proteins 0.000 description 2
- 101700028499 LECG Proteins 0.000 description 2
- 101700063913 LECT Proteins 0.000 description 2
- 239000004472 Lysine Substances 0.000 description 2
- 208000002780 Macular Degeneration Diseases 0.000 description 2
- 208000004451 Membranoproliferative Glomerulonephritis Diseases 0.000 description 2
- 206010061284 Mental disease Diseases 0.000 description 2
- HOKKHZGPKSLGJE-GSVOUGTGSA-N NMDA Chemical compound CN[C@@H](C(O)=O)CC(O)=O HOKKHZGPKSLGJE-GSVOUGTGSA-N 0.000 description 2
- 208000009025 Nervous System Disease Diseases 0.000 description 2
- 206010064062 Neurodevelopmental disease Diseases 0.000 description 2
- 206010029305 Neurological disorder Diseases 0.000 description 2
- 101710034340 Os04g0173800 Proteins 0.000 description 2
- 229920001363 Polidocanol Polymers 0.000 description 2
- 229920001213 Polysorbate 20 Polymers 0.000 description 2
- 102000001708 Protein Isoforms Human genes 0.000 description 2
- 108010029485 Protein Isoforms Proteins 0.000 description 2
- 206010061920 Psychotic disease Diseases 0.000 description 2
- 229920002684 Sepharose Polymers 0.000 description 2
- 210000000278 Spinal Cord Anatomy 0.000 description 2
- 108010090804 Streptavidin Proteins 0.000 description 2
- 210000000225 Synapses Anatomy 0.000 description 2
- 230000004913 activation Effects 0.000 description 2
- 238000007792 addition Methods 0.000 description 2
- 230000001058 adult Effects 0.000 description 2
- 239000000556 agonist Substances 0.000 description 2
- 230000033228 biological regulation Effects 0.000 description 2
- 239000012472 biological sample Substances 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- UIIMBOGNXHQVGW-UHFFFAOYSA-M buffer Substances [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 201000008779 central nervous system disease Diseases 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 230000019771 cognition Effects 0.000 description 2
- 238000010192 crystallographic characterization Methods 0.000 description 2
- 230000004059 degradation Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 2
- VYFYYTLLBUKUHU-UHFFFAOYSA-N dopamine Chemical compound NCCC1=CC=C(O)C(O)=C1 VYFYYTLLBUKUHU-UHFFFAOYSA-N 0.000 description 2
- 239000012149 elution buffer Substances 0.000 description 2
- 230000002708 enhancing Effects 0.000 description 2
- SFNALCNOMXIBKG-UHFFFAOYSA-N ethylene glycol monododecyl ether Chemical compound CCCCCCCCCCCCOCCO SFNALCNOMXIBKG-UHFFFAOYSA-N 0.000 description 2
- 230000007274 generation of a signal involved in cell-cell signaling Effects 0.000 description 2
- 230000000575 glycinergic Effects 0.000 description 2
- 201000001066 hemolytic-uremic syndrome Diseases 0.000 description 2
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 101700036391 lecA Proteins 0.000 description 2
- 239000002523 lectin Substances 0.000 description 2
- 239000003446 ligand Substances 0.000 description 2
- 101700001016 mbhA Proteins 0.000 description 2
- 230000000813 microbial Effects 0.000 description 2
- 201000008895 mood disease Diseases 0.000 description 2
- 210000000653 nervous system Anatomy 0.000 description 2
- 210000002569 neurons Anatomy 0.000 description 2
- 239000002773 nucleotide Substances 0.000 description 2
- 125000003729 nucleotide group Chemical group 0.000 description 2
- ONJQDTZCDSESIW-UHFFFAOYSA-N polidocanol Chemical compound CCCCCCCCCCCCOCCOCCOCCOCCOCCOCCOCCOCCOCCO ONJQDTZCDSESIW-UHFFFAOYSA-N 0.000 description 2
- 229960002226 polidocanol Drugs 0.000 description 2
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000011535 reaction buffer Substances 0.000 description 2
- 239000011347 resin Substances 0.000 description 2
- 229920005989 resin Polymers 0.000 description 2
- 239000012146 running buffer Substances 0.000 description 2
- 238000001542 size-exclusion chromatography Methods 0.000 description 2
- 239000007790 solid phase Substances 0.000 description 2
- 230000000946 synaptic Effects 0.000 description 2
- 230000005062 synaptic transmission Effects 0.000 description 2
- 230000001225 therapeutic Effects 0.000 description 2
- 238000002560 therapeutic procedure Methods 0.000 description 2
- 210000001519 tissues Anatomy 0.000 description 2
- 229920000160 (ribonucleotides)n+m Polymers 0.000 description 1
- AYXZIZMZXAORLO-UFLZEWODSA-N 5-[(3aS,4S,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]pentanoic acid;1-hydroxypyrrolidine-2,5-dione Chemical compound ON1C(=O)CCC1=O.N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 AYXZIZMZXAORLO-UFLZEWODSA-N 0.000 description 1
- LPQOADBMXVRBNX-UHFFFAOYSA-N AC1LDCW0 Chemical compound Cl.C1CN(C)CCN1C1=C(F)C=C2C(=O)C(C(O)=O)=CN3CCSC1=C32 LPQOADBMXVRBNX-UHFFFAOYSA-N 0.000 description 1
- 230000035533 AUC Effects 0.000 description 1
- 108010062271 Acute-Phase Proteins Proteins 0.000 description 1
- 102000011767 Acute-Phase Proteins Human genes 0.000 description 1
- 108010003529 Alternative Pathway Complement C3 Convertase Proteins 0.000 description 1
- MGRVRXRGTBOSHW-UHFFFAOYSA-N Aminomethylphosphonic acid Chemical compound NCP(O)(O)=O MGRVRXRGTBOSHW-UHFFFAOYSA-N 0.000 description 1
- 208000007415 Anhedonia Diseases 0.000 description 1
- 108010047814 Antigen-Antibody Complex Proteins 0.000 description 1
- 229920002395 Aptamer Polymers 0.000 description 1
- 201000006062 Asperger syndrome Diseases 0.000 description 1
- 206010003484 Asperger's disease Diseases 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 208000008581 Brain Disease Diseases 0.000 description 1
- 208000001183 Brain Injury Diseases 0.000 description 1
- 210000000133 Brain Stem Anatomy 0.000 description 1
- 102000025380 C-Reactive Protein Human genes 0.000 description 1
- 108010074051 C-Reactive Protein Proteins 0.000 description 1
- 101710012938 CAGL0K08184g Proteins 0.000 description 1
- 101710042043 CNE03890 Proteins 0.000 description 1
- 102100009686 CXCL9 Human genes 0.000 description 1
- 101700052645 CXCL9 Proteins 0.000 description 1
- 206010007776 Catatonia Diseases 0.000 description 1
- 210000003169 Central Nervous System Anatomy 0.000 description 1
- 206010008129 Cerebral palsy Diseases 0.000 description 1
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 1
- 102000019034 Chemokines Human genes 0.000 description 1
- 108010012236 Chemokines Proteins 0.000 description 1
- 210000000349 Chromosomes Anatomy 0.000 description 1
- 206010057668 Cognitive disease Diseases 0.000 description 1
- 206010010904 Convulsion Diseases 0.000 description 1
- 206010070976 Craniocerebral injury Diseases 0.000 description 1
- 206010012239 Delusion Diseases 0.000 description 1
- 206010061818 Disease progression Diseases 0.000 description 1
- 208000001187 Dyskinesias Diseases 0.000 description 1
- 108010067770 Endopeptidase K Proteins 0.000 description 1
- 210000002744 Extracellular Matrix Anatomy 0.000 description 1
- 101700023515 FABP Proteins 0.000 description 1
- 108009000484 Fragile X Syndrome Proteins 0.000 description 1
- 208000001914 Fragile X Syndrome Diseases 0.000 description 1
- BTCSSZJGUNDROE-UHFFFAOYSA-N GABA Chemical compound NCCCC(O)=O BTCSSZJGUNDROE-UHFFFAOYSA-N 0.000 description 1
- 229960002897 Heparin Drugs 0.000 description 1
- ZFGMDIBRIDKWMY-PASTXAENSA-N Heparin Chemical compound CC(O)=N[C@@H]1[C@@H](O)[C@H](O)[C@@H](COS(O)(=O)=O)O[C@@H]1O[C@@H]1[C@@H](C(O)=O)O[C@@H](O[C@H]2[C@@H]([C@@H](OS(O)(=O)=O)[C@@H](O[C@@H]3[C@@H](OC(O)[C@H](OS(O)(=O)=O)[C@H]3O)C(O)=O)O[C@@H]2O)CS(O)(=O)=O)[C@H](O)[C@H]1O ZFGMDIBRIDKWMY-PASTXAENSA-N 0.000 description 1
- 210000003494 Hepatocytes Anatomy 0.000 description 1
- 229940088597 Hormone Drugs 0.000 description 1
- 102100000995 IBSP Human genes 0.000 description 1
- 229940072221 IMMUNOGLOBULINS Drugs 0.000 description 1
- 210000000987 Immune System Anatomy 0.000 description 1
- 102000018071 Immunoglobulin Fc Fragments Human genes 0.000 description 1
- 108010091135 Immunoglobulin Fc Fragments Proteins 0.000 description 1
- 102000018358 Immunoglobulins Human genes 0.000 description 1
- 108060003951 Immunoglobulins Proteins 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 108010028750 Integrin-Binding Sialoprotein Proteins 0.000 description 1
- 102000004310 Ion Channels Human genes 0.000 description 1
- 108090000862 Ion Channels Proteins 0.000 description 1
- 210000003734 Kidney Anatomy 0.000 description 1
- NRYBAZVQPHGZNS-ZSOCWYAHSA-N Leptin Chemical compound O=C([C@H](CO)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)CNC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](N)CC(C)C)CCSC)N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CS)C(O)=O NRYBAZVQPHGZNS-ZSOCWYAHSA-N 0.000 description 1
- 102000016267 Leptin Human genes 0.000 description 1
- 108010092277 Leptin Proteins 0.000 description 1
- 102000004086 Ligand-Gated Ion Channels Human genes 0.000 description 1
- 108090000543 Ligand-Gated Ion Channels Proteins 0.000 description 1
- 102100014893 MMP3 Human genes 0.000 description 1
- 101700040359 MMP3 Proteins 0.000 description 1
- 229920002521 Macromolecule Polymers 0.000 description 1
- 206010025421 Macule Diseases 0.000 description 1
- 102000009112 Mannose-Binding Lectin Human genes 0.000 description 1
- 108010087870 Mannose-Binding Lectin Proteins 0.000 description 1
- 102000016193 Metabotropic Glutamate Receptors Human genes 0.000 description 1
- 108010010914 Metabotropic Glutamate Receptors Proteins 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 108060004843 Mical Proteins 0.000 description 1
- 208000010428 Muscle Weakness Diseases 0.000 description 1
- 206010028372 Muscular weakness Diseases 0.000 description 1
- 241001182492 Nes Species 0.000 description 1
- 102000005665 Neurotransmitter Transport Proteins Human genes 0.000 description 1
- 108010084810 Neurotransmitter Transport Proteins Proteins 0.000 description 1
- 210000000440 Neutrophils Anatomy 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 229940055729 Papain Drugs 0.000 description 1
- 108090000526 Papain Proteins 0.000 description 1
- 206010033799 Paralysis Diseases 0.000 description 1
- 108010076181 Proinsulin Proteins 0.000 description 1
- 210000004129 Prosencephalon Anatomy 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 239000012614 Q-Sepharose Substances 0.000 description 1
- 206010038428 Renal disease Diseases 0.000 description 1
- 102000004167 Ribonuclease P Human genes 0.000 description 1
- 108090000621 Ribonuclease P Proteins 0.000 description 1
- 241000283984 Rodentia Species 0.000 description 1
- 101700083752 SCR18 Proteins 0.000 description 1
- 101700043450 SCR20 Proteins 0.000 description 1
- 101700080072 SCR4 Proteins 0.000 description 1
- 101700071866 SCR5 Proteins 0.000 description 1
- 206010039911 Seizure Diseases 0.000 description 1
- 206010040030 Sensory loss Diseases 0.000 description 1
- 108010003723 Single-Domain Antibodies Proteins 0.000 description 1
- 101710004918 Smlt3054 Proteins 0.000 description 1
- 206010041466 Speech disease Diseases 0.000 description 1
- 229920001385 Spiegelmer Polymers 0.000 description 1
- 229940035295 Ting Drugs 0.000 description 1
- 208000005765 Traumatic Brain Injury Diseases 0.000 description 1
- 230000036462 Unbound Effects 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 230000001594 aberrant Effects 0.000 description 1
- 230000003213 activating Effects 0.000 description 1
- 229930002945 all-trans-retinaldehyde Natural products 0.000 description 1
- 229960003692 aminobutyric acid Drugs 0.000 description 1
- 230000033115 angiogenesis Effects 0.000 description 1
- 210000004102 animal cell Anatomy 0.000 description 1
- 230000000561 anti-psychotic Effects 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000038129 antigens Human genes 0.000 description 1
- 108091007172 antigens Proteins 0.000 description 1
- 201000007201 aphasia Diseases 0.000 description 1
- 238000003782 apoptosis assay Methods 0.000 description 1
- 238000002820 assay format Methods 0.000 description 1
- 201000007185 autism spectrum disease Diseases 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- 230000003542 behavioural Effects 0.000 description 1
- 239000003150 biochemical marker Substances 0.000 description 1
- 230000031018 biological processes and functions Effects 0.000 description 1
- 239000000091 biomarker candidate Substances 0.000 description 1
- 238000007413 biotinylation Methods 0.000 description 1
- 230000003925 brain function Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- CURLTUGMZLYLDI-UHFFFAOYSA-N carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 1
- 235000011089 carbon dioxide Nutrition 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000001413 cellular Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000003196 chaotropic Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 201000008790 communication disease Diseases 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000001351 cycling Effects 0.000 description 1
- 231100000868 delusion Toxicity 0.000 description 1
- 230000001809 detectable Effects 0.000 description 1
- 239000010432 diamond Substances 0.000 description 1
- 239000012470 diluted sample Substances 0.000 description 1
- 229960003638 dopamine Drugs 0.000 description 1
- 230000003291 dopaminomimetic Effects 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 230000003028 elevating Effects 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 230000002996 emotional Effects 0.000 description 1
- 230000002255 enzymatic Effects 0.000 description 1
- 230000002964 excitative Effects 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 238000005755 formation reaction Methods 0.000 description 1
- 230000003371 gabaergic Effects 0.000 description 1
- 201000002406 genetic disease Diseases 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000000984 immunochemical Effects 0.000 description 1
- 230000001771 impaired Effects 0.000 description 1
- 102000006495 integrins Human genes 0.000 description 1
- 108010044426 integrins Proteins 0.000 description 1
- 201000002199 language disease Diseases 0.000 description 1
- 229940039781 leptin Drugs 0.000 description 1
- 238000007477 logistic regression Methods 0.000 description 1
- BMQVDVJKPMGHDO-UHFFFAOYSA-K magnesium;potassium;chloride;sulfate;trihydrate Chemical compound O.O.O.[Mg+2].[Cl-].[K+].[O-]S([O-])(=O)=O BMQVDVJKPMGHDO-UHFFFAOYSA-K 0.000 description 1
- 230000003340 mental Effects 0.000 description 1
- 230000002503 metabolic Effects 0.000 description 1
- CKJNUZNMWOVDFN-UHFFFAOYSA-N methanone Chemical compound O=[CH-] CKJNUZNMWOVDFN-UHFFFAOYSA-N 0.000 description 1
- 230000000051 modifying Effects 0.000 description 1
- 238000010369 molecular cloning Methods 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 238000002515 oligonucleotide synthesis Methods 0.000 description 1
- 125000001820 oxy group Chemical group [*:1]O[*:2] 0.000 description 1
- 235000019834 papain Nutrition 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 108091008117 polyclonal antibodies Proteins 0.000 description 1
- 238000003752 polymerase chain reaction Methods 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 238000007781 pre-processing Methods 0.000 description 1
- OZAIFHULBGXAKX-UHFFFAOYSA-N precursor Substances N#CC(C)(C)N=NC(C)(C)C#N OZAIFHULBGXAKX-UHFFFAOYSA-N 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 230000005522 programmed cell death Effects 0.000 description 1
- 230000001737 promoting Effects 0.000 description 1
- 238000002731 protein assay Methods 0.000 description 1
- 239000012474 protein marker Substances 0.000 description 1
- 238000011002 quantification Methods 0.000 description 1
- 230000001105 regulatory Effects 0.000 description 1
- 230000002207 retinal Effects 0.000 description 1
- 235000020945 retinal Nutrition 0.000 description 1
- 239000011604 retinal Substances 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000009738 saturating Methods 0.000 description 1
- 230000001953 sensory Effects 0.000 description 1
- SQVRNKJHWKZAKO-OQPLDHBCSA-N sialic acid Chemical compound CC(=O)N[C@@H]1[C@@H](O)C[C@@](O)(C(O)=O)OC1[C@H](O)[C@H](O)CO SQVRNKJHWKZAKO-OQPLDHBCSA-N 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 230000002269 spontaneous Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 201000009032 substance abuse Diseases 0.000 description 1
- 231100000736 substance abuse Toxicity 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000003956 synaptic plasticity Effects 0.000 description 1
- 201000010874 syndrome Diseases 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000002194 synthesizing Effects 0.000 description 1
- 102000002938 thrombospondin family Human genes 0.000 description 1
- 108060008245 thrombospondin family Proteins 0.000 description 1
- 230000007838 tissue remodeling Effects 0.000 description 1
- 231100000730 tolerability Toxicity 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 238000004450 types of analysis Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/76—Chemiluminescence; Bioluminescence
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54366—Apparatus specially adapted for solid-phase testing
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
- G01N33/6896—Neurological disorders, e.g. Alzheimer's disease
Abstract
in vitro method of predicting a response for patients, having neurodevelopmental, neurological or neuropsychiatric disorders, if treated with a compound, targeting the glutamatergic pathway, comprises the steps i) determining the protein concentration of one, two, three, four five or six members of the complement factor H family or a mixture or a combination thereof in a sample of a patient and ii) comparing the protein concentration determined in step i) to a value representative of the protein concentration of one, two three, four, five or six members of complement factor H family in patients, having neurodevelopmental, neurological or neuropsychiatric disorders. A higher protein concentration of one, two three, four five or six members from complement factor H family in the sample of the patient having neurodevelopmental, neurological or neuropsychiatric disorders is indicative for a patient who will derive clinical benefit from this treatment. The method further comprises iii) selecting this treatment for patients having neurodevelopmental, neurological or neuropsychiatric disorders. Also disclosed is an in vitro method of predicting a response for patients, having neurodevelopmental, neurological or neuropsychiatric disorders, if treated with a compound targeting the glutamatergic pathway, wherein the protein concentration of individual members of the complement factor H family or a mixture or a combination thereof are determined by measuring genetic variants of complement factor H family members. Complement factor H family members are useful as predictive markers for patients who are treated with compounds targeting the glutamatergic pathway or for patients who are treated with a glycine reuptake inhibitor. of the complement factor H family or a mixture or a combination thereof in a sample of a patient and ii) comparing the protein concentration determined in step i) to a value representative of the protein concentration of one, two three, four, five or six members of complement factor H family in patients, having neurodevelopmental, neurological or neuropsychiatric disorders. A higher protein concentration of one, two three, four five or six members from complement factor H family in the sample of the patient having neurodevelopmental, neurological or neuropsychiatric disorders is indicative for a patient who will derive clinical benefit from this treatment. The method further comprises iii) selecting this treatment for patients having neurodevelopmental, neurological or neuropsychiatric disorders. Also disclosed is an in vitro method of predicting a response for patients, having neurodevelopmental, neurological or neuropsychiatric disorders, if treated with a compound targeting the glutamatergic pathway, wherein the protein concentration of individual members of the complement factor H family or a mixture or a combination thereof are determined by measuring genetic variants of complement factor H family members. Complement factor H family members are useful as predictive markers for patients who are treated with compounds targeting the glutamatergic pathway or for patients who are treated with a glycine reuptake inhibitor.
Description
Case 27353
A method for predicting clinical benefit in the treatment of neurodevelopmental,
neurological or sychiatric disorders
The t invention relates to methods which are predictive for the clinical benefit for
the treatment of patients with a compound, targeting the glutamatergic pathway, having
evelopmental, neurological or neuropsychiatric disorders.
A neurodevelopmental disorder is an ment of the growth and development of the
brain or central nervous system. A narrower use of the term refers to a disorder of brain on
that affects emotion, learning ability and , and social competence, skills and behaviors
and that unfolds as the individual grows. Disorders considered to be neurodevelopmental in
origin or to have neurodevelopmental consequences when they occur in infancy and childhood
include autism and autism spectrum disorders such as Asperger syndrome, traumatic brain injury
(including ital injuries such as those that cause cerebral palsy, communication, speech
and language disorders, genetic disorders such as fragile-X syndrome and Down me.
Neurodevelopmental disorders are associated with widely varying degrees of mental, emotional,
physical and economic burden to individuals, families and society in general.
A neurological er is a disorder of the body’s nervous system. Structural,
mical or ical abnormalities in the brain, spinal cord, or in the nerves leading to or
from them, can result in symptoms such as paralysis, muscle weakness, poor coordination, loss
of sensation, seizures, confusion, pain, apathy and altered states of consciousness. There are
many recognized neurological disorders, some relatively common, but many rare. The World
Health Organization estimated in 2006 that neurological disorders and their sequelae affect as
many as one billion people ide, and fied health inequalities and social
stigma/discrimination as major factors contributing to the associated disability and suffering.
Neuropsychiatry is the branch of medicine dealing with mental disorders attributable to
diseases of the nervous system and it is also closely related to the field of psychiatry and
behavioral neurology, which is a subspecialty of neurology that addresses clinical problems of
cognition and/or behavior caused by brain injury or brain disease.
Pop/15.06.2012
Neurodevelopmental, neurological or neuropsychiatric disorders include schizophrenia,
r disorders, substance dependence (alcohol, e), autism and obsessive compulsive
disorders (OCD), which are the most red indications with respect to the present invention.
Described herein is the use of the proteins of the complement factor H family as
predictive markers for clinical benefit for patients which are d with a compound, targeting
the glutamatergic y, for example with a glycine reuptake inhibitor. Also described herein
is a method for predicting drug response from a sample, derived from an individual by
measuring proteins of complement factor H family in said sample in vitro.
Complement factor H family means that it may be an dual of CFH proteins
containing CFH, CFHR1, CFHR2, CFHR3, CFHR4A, CFHR4B and CFHR5 or may be a
mixture thereof.
Glycine Reuptake Inhibitors, which targets GlyT1, (GRI) are a novel class of compounds
that are thought to enhance NMDA or (NMDA-R) mediated transmission by elevating
extracellular concentrations of glycine. Evidence from studies in healthy individuals, psychotic
patients and animals as well as from genetic is has accumulated over the past 15 years of
the involvement of NMDA receptor (NMDA-R) hypo-function in the pathophysiology of
neurodevelopmental, neurological or neuropsychiatric disorders.
In CNS, glycine has two major roles for controlling sensory and higher brain on: it
is an inhibitory neurotransmitter in glycinergic neurons and an exitatory neurotransmitter as a co-
agonist with ate of the glutamatergic transmission of the NMDA receptor.
Glutamate is the main excitatory neurotransmitter in the brain and activates NMDA and
non-NMDA receptors (ligand-gated ion channels, i.e. AMPA, kainite and tropic receptors,
i.e. mGluR 1 – 8). NMDA receptors are d on different neurons, such as atergic,
dopaminergic and GABAergic. NMDA receptors can therefore directly affect Glutamate, GABA
and Dopamine.
NMDA receptors are ligands gated ion channels meaning they require for their activation
the binding of both ate and glycine. Glycine acts as modulator of glutamate: it increases
the y of glutamate thus enhancing its effect on activating the receptor. In the developing
brain and, to a lesser extent in the mature brain, NMDA receptors play a crucial role in
synaptogenesis and in promoting synaptic maintenance and stabilization (via increased synaptic
plasticity). NMDA receptors are therefore involved in several brain functions both in mature and
in developing brain.
A dysfunction in glutamatergic neurotransmission is involved in the pathophysiology of
mood disorders and schizophrenia. Thus, it is likely that the NMDA receptor e-modulatory
site is a therapeutic target for improving cognition and reducing ve symptoms in
schizophrenia.
As glycine is an obligatory co-agonist at the NMDA-R complex, one strategy to e
NMDA-R mediated neurotransmission is to elevate extracellular concentrations of e in the
local microenvironment of NMDA receptors. Glycine elevation can be achieved by inhibition of
GRI, which is responsible for e removal from the synaptic cleft. Possible advantages over
the existing ogical and neuropsychiatric therapies include the potential of glycine reuptake
inhibitors in having good efficacy, as well as an improved tolerability profile for the treatment of
negative and positive symptoms in schizophrenia, bipolar disorders, substance dependence
(alcohol, cocaine), autism or obsessive compulsive disorders (OCD).
GlyT1 belongs to the superfamily of neurotransmitter transporters, like SERT, NET or
DAT. GlyT1 nds the atergic synapse in the forebrain. It maintains extracellular
glycine concentration in the synapse below the saturating level at its binding site on NMDA
receptor’s NR1 site. In the caudal brain region and spinal cord, GlyT1 controls both
atergic (via NMDA receptors) and glycinergic transmission.
It is known that glycine reuptake inhibitors may be used for the treatment of
neurodevelopmental, neurological or neuropsychiatric disorders, such as schizophrenia.
A suitable e reuptake inhibitor (GRI) which is used in the content of the present
invention is [4-(3-fluorotrifluoromethyl-pyridinyl)-piperazinyl]-[5-methanesulfonyl
((S)-2,2,2-trifluoromethyl-ethoxy)-phenyl]-methanone.
Schizophrenia is a severe mental disorder typically appearing in late adolescence or early
ood with a word-wide ence of approximately 1% of the adult population which has
enormous social and economic impact. The criteria of the Association of European Psychiatrists
(ICD) and the American Psychiatric Association (DSM) for the diagnosis of schizophrenia
require that two or more characteristic symptoms be present: delusions, inations,
disorganized speech, grossly disorganized or catatonic behavior, or ve symptoms (alogia,
affective flattening, lack of motivation, anhedonia), and that other requirements, such as
excluding affective disorders, and the presence of impaired function, be present. As a group,
people with schizophrenia have functional impairments that may begin in childhood, continue
throughout adult life and make most patients unable to maintain normal employment or
ise have normal social function. [2,3]. They also have a shortened an compared to
the general population, and suffer from an increased prevalence of a wide variety of other
sychiatric syndromes, including serious, substance abuse, obsessive-compulsive
symptoms and abnormal involuntary movements prior to antipsychotic treatment. Schizophrenia
is also associated with a wide range of cognitive impairments, the severity of which limits their
function, even when psychotic ms are well controlled.
Other indications associated with atergic transmission are bipolar disorders,
substance dependence (alcohol, cocaine), autism and obsessive compulsive disorders (OCD).
It has long been acknowledged that there is a need to develop methods of individualizing
treatment of CNS diseases such as schizophrenia, bipolar disorders, substance ence
(alcohol, cocaine), autism and obsessive compulsive disorders (OCD).
With the development of targeted disease treatments, there is a particular interest in
methodologies which could provide a molecular profile of the target, (i.e. those that are
tive for clinical benefit).
Therefore, it is an object of the present invention to provide an in vitro method to t
al benefit to a compound, targeting the atergic pathway, e.g. to identify individuals
who will derive clinical benefit to ent with a compound, ing the glutamatergic
pathway in related diseases, which are schizophrenia, r disorders, nce dependence
(alcohol, cocaine), autism and obsessive compulsive disorders (OCD), or at least to provide the
public with a useful choice.
For this purpose, a predictive marker present in the circulation which is detectable in body
fluids (e.g. blood, serum or ) has to be found.
In order to be of clinical utility, a predictive marker should be able to discriminate those
individuals who will derive clinical benefit from those individuals who will not derive clinical
benefit from treatment with a compound, targeting the glutamatergic pathway. The predictive
sensitivity or specificity of a test is best assessed by its receiver-operating characteristics.
Whole blood, serum or plasma are the most widely used sources of sample in clinical
routine. The identification of a predictive marker that would aid in the identification of
individuals who are likely to respond to treatment with a compound, targeting the glutamatergic
pathway in schizophrenia and other diseases could lead to a method that would greatly aid in the
treatment and in the management of these diseases. Therefore, an urgent clinical need exists to
improve the treatment of es, targeting the glutamatergic y, for example GRI related
diseases.
The object of the present invention is further to investigate whether a biochemical marker
can be fied which may be used in predicting response to a compound, targeting the
glutamatergic pathway, for example to GRI in vitro in a body fluid sample, or at least to provide
the public with a useful choice.
Surprisingly it has been found that an in vitro determination of the concentration of
protein CFH family in a sample allows the prediction of a clinical benefit from the ent
with GRI for patients with neurodevelopmental, neurological or sychiatric disorders.
In this context it was found that determining the protein concentration of ment
factor H family members in the blood, serum or plasma sample of a patient is indicative for a
patient who will derive clinical benefit from the treatment. For this purpose the protein
concentration of complement factor H family members are compared to a value representative of
the protein concentration of ment factor H family members of a population of patients
deriving no clinical benefit from the treatment, n a higher protein concentration of
complement factor H family members in the blood, serum or plasma sample of the patient is
indicative for a patient who will derive clinical benefit from the treatment.
In a first aspect, the present invention provides an in vitro method of predicting a
se for patients, having ve or positive symptoms of schizophrenia, if treated with a
glycine reuptake inhibitor (GRI), comprising the steps:
i) ining the protein concentration of complement factor H related protein 1
(CFHR1) in a sample of a patient,
ii) comparing the protein concentration determined in step i) to a value representative of
the protein tration of complement factor H related n 1 (CFHR1) in patients, having
negative or positive symptoms of schizophrenia,
iii) wherein a higher protein concentration of complement factor H d protein 1
(CFHR1) in the sample of the patient having ve or positive symptoms of schizophrenia is
indicative for a patient who will derive clinical benefit from this treatment and
iv) selecting this treatment for patients having negative or positive symptoms of
schizophrenia.
In one embodiment of this aspect, the protein concentration of complement factor H
related n 1 (CFHR1) is determined by ELISA based technology.
In another embodiment of this , the protein concentration of complement factor H
related protein 1 (CFHR1) is determined by measuring genetic variants of complement factor H
related protein 1 (CFHR1).
In r embodiment of this aspect, the protein concentration of complement factor H
related protein 1 (CFHR1) is determined by measuring of genetic variants of CFHR1, either via
measurement of copy number variations of CFHR1 or by measurement of a SNP as a proxy for
the deletion.
In r embodiment of this aspect, the patient is affected with schizoaffective
disorder.
In r embodiment of this aspect, the glycine reuptake inhibitor is [4-(3-fluoro
trifluormethyl-pyridinyl)-piperazinyl]-[5-methanesulfonyl[[(2S)-1,1,1-trifluoropropan-
2-yl]oxy]phenyl]methanone.
In another aspect, the ion provides the use of an antibody specifically binding to a
complement factor H related n 1 (CFHR1) in a method according to the first aspect of the
invention.
In a further aspect, the invention provides the use of a complement factor H related
protein 1 ) protein as a predictive marker for clinical benefit for patients who are treated
with a glycine reuptake inhibitor.
The invention is as defined in the claims. However, the description which follows also
refers to additional methods and uses outside the scope of the present claims. This description is
ed for its technical information.
A new in vitro method of predicting the response of a patient with neurodevelopmental,
neurological or neuropsychiatric ers is described, wherein such patient is treated with a
compound, targeting the glutamatergic pathway, for example with GRI , which method
ses:
- determining the protein concentration of ment factor H family member(s) in the blood,
serum or plasma sample of a patient and
-comparing the protein concentration of complement factor H family member(s) to a value
representative of the protein concentration of complement factor H family member(s) of a
population of patients ng no clinical benefit from the ent, wherein a higher protein
concentration of complement factor H family member(s) in the blood, serum or plasma sample
of the patient is indicative for a patient who will derive clinical benefit from the treatment.
Also described herein is the use of the protein complement factor H family member(s) in
the in vitro assessment of certain CNS diseases, such as schizophrenia, in a sample, wherein a
concentration of protein complement factor H family member(s) above a reference concentration
is indicative for al benefit due to drug treatment of said diseases.
The ment factor H was selected from a panel of 189 proteins (Rules Based
Medicine, Human Discovery Map v 1.0). Baseline and ent serum samples from
schizophrenia patients treated with GRI were analyzed on the Luminex based multiplex ELISA
Human DiscoveryMap® v 1.0.
DiscoveryMAP® is a hensive quantitative assay test, containing 189
protein analytes. It represents the culmination of 10 years of assay development for nes,
chemokines, metabolic markers, hormones, growth factors, tissue remodeling proteins,
angiogenesis markers, acute phase reactants, cancer markers, kidney toxicity markers, CNS
biomarkers and other important serum proteins.
Complement factor H was found superior to the other 189 protein analytes in view of the
following advantages:
- complement factor H shows a strong discrimination n responders and non-responders in
the treatment group
- complement factor H shows no prognostic effect in the placebo group;
- complement factor H serum concentration is not influenced by treatment with GRI within 8
weeks
- complement factor H serum concentration is stable over time.
Complement factor H, also known as factor H, is a sialic acid containing glycoprotein that plays
an integral role in the regulation of the complement-mediated immune system that is involved in
microbial defense, immune complex processing, programmed cell death and age-related macula
degeneration. Complement factor H is the best characterized member of the complement factor
H protein family. The complement factor H family consists of the following members:
complement factor H (CFH), complement factor H-related protein 1 (CFHR1), complement
factor ted protein 2 (CFHR2), complement factor H-related n 3 (CFHR3),
complement factor H-related protein 4 with isoforms 4A and 4B (CFHR4A and ),
complement factor H-related protein 5 (CFHR5). The complement factor H-related proteins are
encoded ream of the complement factor H gene and share a high tration of
homology with subdomains of complement factor H. Complement factor H related proteins share
also functional similarities [4].
The ment system consists of ~40 proteins that are present in body fluids or on cell
and tissue surfaces and is activated in a cascade-like manner by three major pathways [1]. The
alternative pathway is activated continuously at a low rate by the spontaneous hydrolysis of the
central ent C3, the lectin pathway is initiated by mannose binding lectin or ficolins that
ize microbial ydrates and the classical pathway is activated by binding of C1q to
antigen bound immunoglobulins. Enzymatic steps generate active fragments of complement
components and trigger further ication. The three pathways merge at the concentration of
C3, which on activation, is d into C3a and C3b.
Complement factor H protects host cells from injury ing from unrestrained complement
activation. Complement factor H tes complement activation on self cells by possessing
both or activity for the Factor I mediated C3b cleavage, and decay accelerating activity
t the alternative pathway C3 convertase, C3bBb.
Complement factor H protects self cells from complement activation but not
bacteria/viruses. Due to the central role that Complement factor H plays in the regulation of
complement, there are many clinical implications arising from aberrant CFH activity. Mutations
in the Complement factor H gene are ated with severe and diverse diseases including the
rare renal disorders hemolytic uremic syndrome (HUS) and membranoproliferative
glomerulonephritis (MPGN) also termed dense deposit disease (DDD), membranoproliferative
glomuleronephritis type II or dense deposit disease, as well as the more frequent retinal e
age related macular degeneration (AMD). In addition to its complement regulatory activities,
complement factor H has multiple physiological activities and 1) acts as an extracellular matrix
component, 2) binds to cellular receptors of the integrin type, and 3) interacts with a wide
selection of ligands, such as the C-reactive protein, thrombospondin, bone sialoprotein,
ontin, and heparin.
Complement factor H d protein 1 (CFHR1) is a 43 kDa, secreted member of the
factor H family of glycoproteins. It is produced by hepatocytes and circulates as two
differentially glycosylated isoforms (37 kDa and 43 kDa). Mature human CFHR1 is 312 aa in
length. It contains five imately 60 aa SCRs (short consensus repeats/CCPs/SUSHI
repeats) that basically constitute the entire molecule. CFHR1 may play a role in otein
complexes that bind LPS to neutrophils. There is no reported rodent rpart to CFHR1. Over
aa 19143 of the CFHR1 precursor, human CFHR2 and CFHR5 show 98% and 86% aa identity,
respectively.
As used herein
The term “biomarker or marker” means a substance used as an indicator of a biological
state, i.e. of biological processes or pharmacologic response to a therapeutic interaction. A
biomarker or marker indicates a change in expression or state of a protein that correlates with the
risk or progression of a disease, or with the susceptibility of the disease to a given ent.
The term “gene” means a piece of DNA in the host organism.
Gene expression is the process in which information from a gene is used in the synthesis
of a functional gene product, e.g. a protein. The term “expression tration” means the
concentration at which a particular gene is expressed within a cell, tissue or sm. The term
ssion concentration” in context with proteins reflects the amount of a particular protein
present in a cell at a certain time.
The term “protein” means a functional gene product that has been synthesized from the
gene.
The term “a value representative of a protein concentration of complement factor H
family members of a population of patients deriving no clinical benefit from the treatment”
refers to an estimate of a mean expression concentration of a population of patients who do not
derive a al benefit from the treatment. Clinical benefit was d as having a measurable
response compared to o after ≥ 8 weeks.
The term “sample” or "test sample" as used herein refers to a biological sample obtained
from an individual for the purpose of evaluation in vitro. In the methods of the present invention,
the sample or patient sample may comprise in an embodiment of the present invention any body
fluid. Preferred samples are body , such as blood, serum or plasma, with serum or plasma
being most preferred.
The term “mixture” refers to a protein mixture made up by two or more ns that are
simultaneously detected by one protein assay, e.g. a sandwich assay. The simultaneous ion
can be due to similar es being t on the proteins detected by the antibodies used in
the sandwich assay.
The term “combination” relates to independent measurements of two or more protein
analytes measured out of a larger group of proteins, where the order does not matter. The
independent measurement results are combined mathematically, e.g. by calculating a ratio of two
measurement s.
Protein concentrations of complement factor H family member(s), particularly soluble
forms of protein concentrations of complement factor H family member(s) (CFH, CFHR1,
CFHR2, CFHR3, CFHR4A, CFHR4B and CFHR5 or mixtures thereof), are determined in vitro
in an appropriate sample. Preferably, the sample is derived from a human subject, e.g. a
phrenia patient or a person in risk of schizophrenia or a person suspected of having
schizophrenia. Protein concentrations of ment factor H family member(s) are determined
in a blood, serum or plasma sample.
Described herein is a method of ting a response for patients, having neurodevelopmental,
neurological or neuropsychiatric disorders, if treated with a compound, targeting the
glutamatergic pathway, comprising the steps
i) determining the protein concentration of one, two, three, four five or six members of the
complement factor H family or a e or a combination thereof in a sample of a patient,
ii) comparing the protein concentration determined in step i) to a value representative of the
protein concentration of one, two three, four, five or six members of complement factor H family
in patients, having neurodevelopmental, neurological or neuropsychiatric disorders,
iii) wherein a higher protein tration of one, two three, four five or six members from
complement factor H family in the sample of the patient having neurodevelopmental,
neurological or neuropsychiatric disorders is indicative for a patient who will derive clinical
benefit from this treatment and
iv) selecting this treatment for patients having neurodevelopmental, neurological or
neuropsychiatric disorders.
Also described herein is an in vitro method of predicting a response for patients, having
neurodevelopmental, neurological or neuropsychiatric disorders, if treated with a glycine
reuptake inhibitor (GRI), comprising the steps
i) ining the protein tration of one, two, three, four five or six members of the
complement factor H family or a e or a combination thereof in a sample of a patient,
ii) comparing the protein concentration determined in step i) to a value representative of the
protein concentration of one, two three, four, five or six s of complement factor H family
in patients, having neurodevelopmental, neurological or neuropsychiatric disorders,
iii) wherein a higher protein concentration of one, two three, four five or six members from
complement factor H family in the sample of the patient having neurodevelopmental,
neurological or neuropsychiatric disorders is indicative for a patient who will derive clinical
benefit from treatment with GRI, and
iv) selecting GRI treatment for patients having neurodevelopmental, neurological or
sychiatric disorders.
The complement factor H family members include proteins of complement factor H
(CFH), complement factor H related protein 1 (CFHR1), complement factor H related protein 2
(CFHR2), complement factor H related protein 3 (CFHR3), complement factor H d protein
4A CFHR4A), complement factor H related protein 4B (CFHR4B), and ment factor H
related protein 5 (CFHR5).
One embodiment of the method is complement factor H family members as mentioned
above or a mixture or a ation thereof.
One ment of the method is that the ment factor H family members are
complement factor H related protein 1, or a mixture of complement factor H related protein 1
and complement factor H.
One further embodiment is an in vitro method, wherein the complement factor H family
member is complement factor H related protein 1.
One further ment is an in vitro method, wherein the complement factor H family
member is a combination of complement factor H related n 1 and complement factor H
One particularly preferred embodiment is an in vitro method, n the complement
factor H family member is the ratio of complement factor H related protein 1 and complement
factor H.
One embodiment is further an in vitro method, wherein the complement factor H family
member is a combination of complement factor H related protein 3 and complement factor H.
One embodiment is further an in vitro method, wherein the complement factor H family
member is the ratio of complement factor H related protein 3 and complement factor H.
One embodiment is further an in vitro method, wherein the complement factor H family
member is a combination of either complement factor H related protein 3 or complement factor
H related protein 1 and one of the following complement factors: complement factor H related
protein 2, complement factor H related protein 4A , complement factor H related protein
4B,complement factor H related protein 5 and complement factor H.
One embodiment is further an in vitro method, wherein the complement factor H family
member is the ratio of either complement factor H related protein 3 or complement factor H
d protein 1 and one of the following complement s: complement factor H related
protein 2, complement factor H related protein 4A , complement factor H d protein
4B,complement factor H related protein 5 and complement factor H.
The method of predicting a response for patients, having neurodevelopmental, neurological
or neuropsychiatric disorders include ve or positive symptoms of schizophrenia, r
disorder, substance dependence, autism and compulsive disorders.
One ment is a method where the patient is affected with schizoaffective er.
One further embodiment is that the protein concentration of individual members of the
complement factor H family or a mixture or a combination thereof are determined by ELISA
based logy.
Also described herein is an in vitro method of predicting a response for patients, having
neurodevelopmental, ogical or neuropsychiatric disorders, if treated with a compound,
targeting the glutamatergic pathway which may be a glycine reuptake inhibitor (GRI), wherein
the n concentration of individual members of the ment factor H family or a mixture
or a combination thereof are determined by measuring genetic variants of complement factor H
family members [5].
Also described herein is an in vitro method of predicting a response for patients, having
neurodevelopmental, neurological or neuropsychiatric ers, if treated with a glycine
reuptake inhibitor (GRI) wherein the protein concentration of CFHR1 is determined by
measuring of genetic variants of CFHR1, either via measurement of copy number ions of
CFHR1 or by measurement of a SNP as a proxy for the deletion.
Also bed herein is the use of an antibody specifically binding to a protein of the
complement factor H family.
The in vitro method comprises the use of a GRI, which is [4-(3-fluorotrifluormethylpyridinyl
razinyl]-[5-methanesulfonyl[[(2S)-1,1,1-trifluoropropan
yl]oxy]phenyl]methanone.
Also described herein is a complement factor H family member for use as a predictive
marker for patients who are treated with a compound targeting the glutamatergic pathway, for
example with a glycine ke inhibitor (GRI).
Also described herein is the use of complement factor H family member as a predictive
marker for clinical benefit for ts, having neurodevelopmental, ogical or
neuropsychiatric disorders, if treated with a compound targeting the glutamatergic pathway, for
example with a glycine reuptake tor (GRI).
Also described herein is a complement factor H family member as a predictive marker for
clinical benefit for patients, having neurodevelopmental, neurological or neuropsychiatric
disorders, if treated with a compound targeting the glutamatergic pathway, for example with a
e reuptake inhibitor (GRI).
The term "reference sample" as used herein refers to a biological sample provided from a
reference group of patients deriving no clinical benefit from the treatment for the purpose of
evaluation in vitro. The term "reference concentration" as used herein refers to a value
established in a reference group of patients deriving no clinical benefit from the treatment.
It is known to a person skilled in the art that the measurement results of step (i) according
to the method(s) of the present invention will be ed to a reference concentration. Such
reference tration can be determined using a negative nce sample, a positive
reference sample, or a mixed nce sample comprising one or more than one of these types of
controls.
The expression "comparing the concentration determined to a reference concentration” is
merely used to further illustrate what is obvious to the skilled artisan anyway. A reference
concentration is established in a control sample. The l sample may be an internal or an
external control sample. In one embodiment an internal control sample is used, i.e. the marker
concentration(s) is(are) ed in the test sample as well as in one or more other sample(s)
taken from the same subject to determine if there are any changes in the concentration(s) of said
marker(s). In another embodiment an al control sample is used. For an external control
sample the presence or amount of a marker in a sample derived from the individual is compared
to its presence or amount in an individual known to suffer from, or known to be at risk of, a
given condition; or an individual known to be free of a given condition, i.e., "normal individual".
For example, a marker concentration in a t sample can be ed to a concentration
known to be associated with a specific course of evelopmental, neurological or
neuropsychiatric disorders.
Usually the sample’s marker concentration is directly or indirectly correlated with a
diagnosis and the marker concentration is e.g. used to determine whether an individual is at risk
for neurodevelopmental, neurological or neuropsychiatric disorders.
atively, the sample’s marker concentration can e.g. be compared to a marker
concentration known to be associated with a response to therapy in schizophrenia patients, the
diagnosis of schizophrenia, the guidance for selecting an appropriate drug to schizophrenia, in
judging the risk of disease progression, or in the follow-up of schizophrenia patients. Depending
on the intended diagnostic use an appropriate control sample is chosen and a control or reference
value for the marker established therein. It will be appreciated by the d artisan that such
l sample in one embodiment is obtained from a reference population that is age-matched
and free of nding diseases. As also clear to the skilled artisan, the te marker values
established in a control sample will be dependent on the assay used. Preferably samples from
100 well-characterized individuals from the appropriate reference population are used to
establish a reference value. Also preferred the reference population may be chosen to consist of
, 30, 50, 200, 500 or 1000 individuals. Schizophrenia patients who derive no clinical benefit
from GRI treatment ent a preferred reference population for establishing a reference value.
Schizophrenia patients who derive no al benefit from GRI ent can be defined
as patients who show no or only minimal improvement in their symptoms after treatment. In an
embodiment, no or minimal improvement in symptoms is d as less than 20% change in the
Positive and Negative Symptom Scale (PANSS). In another embodiment, no or minimal
improvement in symptoms is defined as worse symptoms, no change or minimally improved
symptoms according to the al Global Impression scale (CGI).
Schizophrenia patients with a specific genotype of CFH family genes represent another
preferred reference population for establishing a control value. In an embodiment, schizophrenia
patients with a homozygous deletion of the CFHR1 and CFHR3 genes represent a preferred
reference population for establishing a nce value. In another embodiment, schizophrenia
patients with a homozygous or heterozygous on of the CFHR1 and CFHR3 genes represent
a preferred nce population for establishing a reference value.
The term “measurement”, "measuring" or "determining" preferably comprises a
qualitative, a semi-quantitative or a quantitative ement. In the present invention a protein
of Complement factor H family members or a mixture f is ed in a body fluid
sample. In a preferred embodiment the measurement is a semi-quantitative measurement, i.e. it is
determined whether the concentration of a protein of Complement factor H family members or a
mixture thereof is above or below a cut-off value.
The values for protein of Complement factor H family s or a mixture thereof as
determined in a control group or a control population are for example used to establish a f
value or a reference range. A value above such cut-off value is considered as indicative for the
prediction of clinical benefit in the treatment of the neurodevelopmental, ogical or
neuropsychiatric disorders.
In an embodiment a fixed cut-off value is established. Such cut-off value is chosen to
match the diagnostic question of interest.
In an embodiment the cut-off is set to result in a specificity of 90%, also preferred the
f is set to result in a specificity of 95%, or also preferred the cut-off is set to result in a
specificity of 98%.
In an embodiment the f is set to result in a sensitivity of 90%, also preferred the
cut-off is set to result in a sensitivity of 95%, or also preferred the cut-off is set to result in a
sensitivity of 98%.
In one embodiment values for a protein of the ment factor H family as determined
in a control group or a control population are used to ish a reference range. In a preferred
embodiment a concentration of a protein of the complement factor H family is considered as
elevated if the value determined is above the 90%-percentile of the reference range. In further
preferred embodiments a protein concentration of a complement factor H family member is
considered as elevated if the value determined is above the 95%-percentile, the 96%-percentile,
the 97%-percentile or the 97.5%-percentile of the reference range.
A value above the cut-off value can for example be indicative for an increased clinical
benefit in the treatment of the neurodevelopmental, neurological or neuropsychiatric disorders.
In a further preferred ment the measurement of a protein of a complement factor
H family (s) is a quantitative measurement. In further embodiments the concentration of
a protein of complement factor H family member(s) is correlated to an increased clinical benefit
in the ent of the neurodevelopmental, neurological or sychiatric disorders.
As the skilled artisan will appreciate, any such assessment is made in vitro. The sample
(test sample) is discarded afterwards. The sample is solely used for the in vitro stic
method of the ion and the al of the sample is not transferred back into the patient’s
body. Typically, the sample is a body fluid sample, e.g., serum, plasma, or whole blood.
The method according to the present invention is based on a liquid or body fluid sample which is
obtained from an individual and on the in vitro determination of protein concentration of
Complement factor H family members or a mixture thereof in such sample. An "individual" as
used herein refers to a single human.
Preferably the protein tration of complement factor H family member(s) are
specifically determined in vitro from a liquid sample by use of a specific binding agent.
Unless defined otherwise, technical and scientific terms used herein have the same
meaning as commonly understood by one of ordinary skill in the art to which this invention
belongs.
Singleton, et al., Dictionary of Microbiology and Molecular Biology, 2nd ed., John Wiley &
Sons, New York, N.Y. (1994);
March, Advanced Organic Chemistry Reactions, Mechanisms and Structure, 4th ed., John Wiley
& Sons, New York, N.Y. (1992);
Lewin, B., Genes V, published by Oxford University Press (1994), ISBN 0854287 9;
Kendrew, J., et al., (eds.), The Encyclopedia of Molecular Biology, published by ell
Science Ltd. (1994), ISBN 02182-9; and
Meyers, R.A., (ed.), lar y and Biotechnology: a Comprehensive Desk Reference,
published by VCH Publishers, Inc. (1995), ISBN 1569 8)
provide one skilled in the art with a general guide to many of the terms used in the present
application.
Techniques for the detection of protein expression of the tive genes described by this
invention include, but are not limited to enzyme linked immunosorbent assay (ELISA).
The practicing of the present invention will employ, unless otherwise indicated,
conventional techniques of lar biology (including recombinant techniques), microbiology,
cell biology, biochemistry, and immunology, which are within the skill of the art. Such
techniques are explained fully in the ture, such as
Sambrook, et al., Molecular Cloning: A Laboratory Manual, second edition, (1989);
Gait, M.J., (ed.) Oligonucleotide Synthesis (1984);
Freshney, R.I., (ed.), Animal Cell Culture (1987); Methods in Enzymology (Academic Press,
Inc.);
Ausubel, F.M., et al., (eds.), Current Protocols in Molecular Biology (1987) and ic updates;
Mullis, et al., (eds.) PCR: The Polymerase Chain Reaction (1994).
The method according to the present invention is based on a liquid or body fluid sample which is
obtained from an individual and on the in vitro determination of protein concentration of
complement factor H family member(s) in such sample. An "individual" as used herein refers to
a single human.
In a preferred embodiment according to the present ion, the protein trations
of complement factor H family member(s) are determined. In an embodiment, the protein
concentration of complement factor H family member(s) are specifically determined in vitro
from a sample by use of a ic g agent.
A specific binding agent is, e.g., a receptor for the complement factor H family
member(s), a lectin binding to complement factor H family (s), an antibody to
complement factor H family member(s), peptide-bodies to complement factor H family
member(s), bi-specific dual binders or bi-specific antibody formats. A specific binding agent has
at least an affinity of 107 l/mol for its corresponding target molecule. The specific binding agent
preferably has an affinity of 108 l/mol or also preferred of 109 l/mol for its target molecule.
As the skilled artisan will appreciate the term specific is used to indicate that other
ecules present in the sample do not significantly bind to the binding agent ic for the
complement factor H protein sequence of SEQ ID NO:1 or complement factor H related proteins
1-5 of SEQ ID NO: 2-7. Preferably, the tration of binding to a ecule other than the
target molecule results in a binding affinity which is at most only 10% or less, only 5% or less
only 2% or less or only 1% or less of the affinity to the target le, respectively. A preferred
specific g agent will fulfil both the above minimum ia for affinity as well as for
specificity.
Examples of specific binding agents are peptides, peptide mimetics, aptamers,
spiegelmers, darpins, ankyrin repeat proteins, Kunitz type domains, antibodies, single domain
antibodies [6] and monovalent fragments of antibodies.
In certain preferred embodiments the specific binding agent is a polypeptide.
In certain red embodiments the specific g agent is an antibody or a monovalent
antibody fragment, preferably a monovalent fragment derived from a monoclonal antibody.
The term "antibody" herein is used in the broadest sense and ically covers
monoclonal antibodies, polyclonal antibodies, multi-specific antibodies (e.g. bi-specific
antibodies) formed from at least two intact antibodies, and antibody fragments so long as they
t the desired biological activity. In certain preferred ments the specific binding
agent is an antibody or a monovalent antibody fragment, preferably a monovalent fragment
derived from a monoclonal antibody.
A specific binding agent preferably is an antibody or a set of antibodies reactive with a
member of the complement factor H family or a combination thereof, e.g. complement factor H
(SEQ ID NO: 1), complement factor H d protein 1 (SEQ ID NO: 2), complement factor H
related protein 2 (SEQ ID NO: 3), complement factor H related protein 3 (SEQ ID NO: 4),
complement factor H related protein 4A (SEQ ID NO: 5), complement factor H related n
4B (SEQ ID NO: 6), complement factor H related protein 5 (SEQ ID NO: 7), either a linear
epitope or a conformational epitope.
As the skilled artisan will appreciate now, that protein trations of complement factor
H family members have been identified as marker which is useful in the assessment of clinical
benefit for the treatment of neurodevelopmental, neurological or neuropsychiatric ers.
For determination of protein concentrations of complement factor H family member(s)
the sample obtained from an dual is incubated in vitro with the specific binding agent for
complement factor H family member(s) under conditions appropriate for formation of a complex
between binding agent and complement factor H family member(s). Such conditions need not be
specified, since the skilled artisan without any inventive effort can easily identify such
appropriate incubation conditions. The amount of complex between binding agent and
complement factor H family (s) is determined for e in an enzyme-linked
immunoassay (ELISA) as described above [7].
Immunoassays are well known to the skilled artisan. Methods for carrying out such assays
as well as practical applications and procedures are summarized in related textbooks. Examples
of related textbooks are
Tijssen, P., Preparation of enzyme-antibody or other enzyme-macromolecule conjugates,
ce and theory of enzyme assays, pp. 221-278,
and various volumes of Methods in Enzymology, Colowick, S.P., and Caplan, N.O. ,
Academic , dealing with immunological ion methods, especially volumes 70, 73, 74,
84, 92 and 121.
Using this assay format, it has been shown that samples from patients can be used to assess
the clinical benefit for patients by treatment. Results are shown in the example section of this
application.
Example
The ion will now be further described in the example below, which is intended as an
illustration only and does not limit the scope of the invention.
Patients and Study Design of Phase 2 GRI Schizophrenia trial
A Phase II study was performed in adults with schizophrenia with inantly
negative symptoms. The individuals were clinically stable and on stable treatment with second
generation ychotics. The study was conducted at le sites in , France, Germany,
Hungary, Japan, Mexico, , Russia, and the US according to ICH Guidelines for Good
Clinical Practice (ClinicalTrials.gov fier: NCT00616798). The study protocol was
approved by the health authorities of each country and the respective ethic committees of each
site. All 323 patients had provided written consent to participate in the study and 224 patients
had consented to the collection and use of serum s for exploratory biomarker analysis.
After a screening and a 4-week run-in period, patients were randomized to double-blind
adjunctive treatment with placebo, 10 mg, 30 mg, or 60 mg of GRI given once a day for a
duration of 8 weeks, followed by 4 week follow-up period. Similar percentages of patients in the
four treatment arms completed the study.
Outcome measures of Phase 2 GRI Schizophrenia trial
Patients were assessed at screening, week -2, baseline, weeks 1, 2, 4, 6, 8, 10 and 12.
ments included the PANSS, Clinical Global Impression - Severity of Illness (CGI-S) and
Clinical Global Impression – Global Improvement ) for overall psychopathology56, CGI-
S and CGI-I of Negative Symptoms (CGI-S-N, CGI-I-N) for assessment of negative symptoms
only57, and standard examinations of extrapyramidal ms (BAS58, SAS59, AIMS60).
The primary efficacy variable ing effects on negative symptoms was the change from
baseline in the PANSS negative m factor score64. Secondary efficacy variables were i)
proportion of patients showing a clinical response in negative symptom defined as a ≥ 20%
change from baseline in the PANSS negative symptom factor score and ii) CGI-I-N. Additional
analyses included change in PANSS total score, remaining PANSS factor scores64 CGI-S and
CGI-I.
Preparation of serum samples
Serum samples were collected at baseline and at 8 weeks of treatment from patients that
had given written consent to the tion and use of serum samples for exploratory biomarker
analysis. Samples were collected in a plain tube t EDTA and allowed to stand for
approximately 30 min or until clotted at room temperatures. Within 60 min of blood collection
s were transferred into a centrifuge and spun at 1500 g for 10 min at 4°C (or at room
temperature if refrigerated centrifuge was not available). Immediately after centrifugation
supernatant (i.e. serum) was transferred into a fresh pre-labelled tube and stored at -70 degree.
ination and results of concentration of CFH from patients from Phase 2
GRI Schizophrenia trial
Baseline serum samples were ed on the Luminex based multiplex ELISA Human
Discovery Map® v 1.0 provided by Rules Based Medicine (Austin Texas). Frozen serum-
samples from the Phase II study were randomized, blinded and sent on dry ice to Rules Based
Medicine for analysis of 189 protein analytes. Protein biomarker measurements were performed
in one batch and results were reported in measurement units (eg ng/ml).
Measurement values were reported if they were within the rd calibration curve.
Values outside the calibration curve were imputed using the calibration limits. Of the 189
measured protein analytes, 26 reported ement values below the lowest calibrator for more
than 70% of the patient samples. These analytes were removed from is. Measurements for
2 related es (pro-insulin and insulin) were aggregated by averaging since they showed
highly correlated measurement values. After pre-processing 162 protein analytes remained in the
analysis data set.
aphics, ne psychopathology and functioning of the biomarker subgroup
versus the total study cohort of Phase 2 GRI Schizophrenia trial
224 out of 323 patients from the Phase 2 GRI phrenia trial had consented to the collection
and use of serum samples for exploratory biomarker analysis. The biomarker subgroup is
representative for the whole study cohort.
Identification of biomarker candidate for response prediction
In the clinical trial as a whole, the primary analysis endpoint was change from baseline in
the PANSS negative ms factor score [8]. The response variable for this analysis was the
change in PANSS Negative Symptom Factor Score between baseline and week 8. For
identification of a biomarker for response prediction protein analytes as measured by RBM were
analyzed as s: The only explanatory variable for this analysis was the baseline
measurement of an individual protein and the analysis was performed on the pooled 10 mg and
mg biomarker population using PP (per protocol) population. For each protein marker
candidate the null hypothesis that change in the PANSS is uncorrelated to the protein
measurement was tested using R^2 of a robust linear model as test statistic. The null hypothesis
was tested for all assayed protein markers in parallel with tion for multiple testing. As
TABLE 1 shows an analyte called “Complement factor H” ranked as the best performing
marker, however, after adjustment for multiple g it did not yield statistical significance.
As a ary nt in the clinical trial the Clinical Global Impression (CGI)
improvement of negative symptoms between baseline and week 8 evaluation was used. This
outcome measure was treated as a binary variable. The positive group consisted of the “Very
much improved” and “Much improved” scores, the negative group consisted of “Minimally
improved”, “No change”, “Minimally worse”, “Much worse” and “Very much . The
analysis was performed on the combined placebo, 10 mg and 30 mg biomarker population using
the per ol (PP) population. atory variables for this is are the baseline
measurements of all measured proteins, the treatment (placebo or pooled 10mg/30mg treatment)
and the treatment-protein interaction. A logistic regression model was used as predictor.
Based on this analysis markers were ranked by their y to discriminate patients with
improvement from patients without improvement in CGI on the 10 and 30 mg treatment arms.
The analyte called “Complement factor H” within the RBM panel was identified as the
only marker reaching tical significance after correction for multiple g with an adjusted
p value of 0.006 (see Table 1).
The ability of serum “complement factor H” baseline levels to predict clinical effect was
independent of demographic and clinical covariates (including baseline PANSS) and there was
no significant difference in “complement factor H" baseline serum concentration between the
different treatment arms. “Complement factor H” showed no association with se in the
placebo group (PANSS or CGI-I).
Further characterization of the antibodies used in the RBM assay revealed that both the
capture and ion antibodies bind within the last 3 short consensus repeats (SCRs) of CFH.
This region is highly homologous to complement factor H related protein 1 (CFHR1: SCR18,
SCR19 and SCR20 share 100%, 100% and 97% identity with SCR 3, SCR4 and SCR5 of
CFHR1, respectively [4].
Human CFH and CFHR1 proteins were isolated from serum and purified. Characterization
of both proteins in the RBM assay showed that CFHR1 was recognized with a 10 fold preference
for CFHR1 over CFH.
Antibodies used for the CFHR1-specific assay
For the development of a specific assay the following monoclonal antibodies were
used: MAB<CFH/CFHR1>M-L20/3 (provider: Thermo Scientific, cat. no.: GAU 02002)
and MAB<CFHR1>M- 442127 (provider: R&D-systems, cat.-no.: 7).
Biotinylation of monoclonal L20/3 Fab-fragments, stoichiometry 1:1.3
Monoclonal mouse IgG (clone: L20/3; er: Thermo Scientific, cat. no.: GAU 020
02) is digested using Papain (3mU/mg IgG) to produce Fab-fragments. Digested Fc-fragments
are eliminated by chromatography on DAE-Sepharose. Purification of Fab by Fcγ-adsorption of
remaining Fc followed by Superdex 200 size exclusion.
To a solution of 10 mg/ml L20/3-Fab fragments in 100 mM KPO4, pH 8.5 50 ul Biotin -N-
hydroxysuccinimide (3.6 mg/ml in DMSO) are added per ml. After 45 min at room temperature,
the sample is dialysed against 100 mM KPO4, 150 mM NaCl, pH 7.2 and frozen.
Ruthenylation of monoclonal 442127 IgG, iometry 1:3
To a on of 5 mg/ml monoclonal mouse IgG (clone 442127, provider: stems,
cat.-no.: MAB4247) in 100 mM KPO4, pH 8.5, 125 ug Ruthenium-(bpy)2-bpyCO-Osu are
. After 75 min at room temperature, ruthenylation is stopped by addition of 10 mM
Lysine. For separation of aggregates riate fractions of sample are collected from Superdex
200 size exclusion chromatography.
Purification of CFHR1 from human serum
For use as calibrator material, native CFHR1 was purified from human serum by
immunoadsorption using the monoclonal antibody L20/3 specific for CFH and CFHR1
according to following procedure: MAB L20/3<CFH,CFHR1>M-IgG-Spherosil resin is used for
purification of CFHR1 from human serum. uted (1:4 in 50 mM TrisHCl, 150 mM NaCl,
pH7.5) serum is passed through Sartoclean CA (0.8µm) and Sartobran P ) filter caps.
Pretreated serum, loaded on the column is washed (10mM TrisHCl, 500 mM NaCl, 0.05%
Tween20, pH 7.5) followed by Q-sepharose running buffer and eluted in gentle elution buffer
(Thermo Scientific) to avoid degradation of analyte and adsorber. For separation of co-bound
CFHR1 and CFH, the immunoadsorber-eluate is run over a MonoQ column for size dependent
elution. The eluate containing CFHR1 as well as ified protein running at different height
in SDS-PAGE analysis is further separated for isoelectricity on a MonoS column resulting in
fractions of pure CFHR1. Pure CFHR1 is applied as calibrator al for the determination of
CFHR1 in the Elecsys-ECLIA test for serum CFHR1 .
Measurement of CFHR1 in human serum or plasma samples using the ECLIA
immunoassay
ECLIA immunoassay for CFHR1 is developed for the specific ement of CFHR1 in
human serum or plasma samples using the Elecsys® Analyzer.
The following bes the assay procedure for the determination of CFHR1 using the
Elecsys® er.
The s CFHR1 immunoassay is an electrochemiluminescence immunoassay (ECLIA)
that functions via the sandwich principle. There are two antibodies included in the assay – a
biotinylated Fab fragment of monoclonal antibody L20/3-Bi (capture dy) and a
ruthenylated monoclonal anti-CFHR1 dy 442127-Ru (detection antibody) – which form
sandwich immunoassay complexes with CFHR1 in the . The complexes are then bound to
solid-phase streptavidin-coated microparticles. The microparticles are magnetically captured
onto the surface of an electrode, and the application of a voltage to the electrode induces
chemiluminescent emission, which is measured by a photomultiplier for readouts. Results are
determined via an instrument-specific calibration curve.
Samples are automatically diluted 1:400 in Diluent (Roche Diluent MultiAssay for Elecsys
Ref. No. 03609987-190). Assay protocol 3 is d allowing 9 min preincubation of 10 ul
diluted sample with 80 ul of ruthenylated <CFHR1>-442127 at 0.5 ug/ml in reaction buffer
(Hepes50 mM, NaCl 150 mM; Thesit/Polidocanol 0.1%; EDTA 1mM; bovine serum albumin
0.5%) before 80 ul of ylated <CFH,CFHR1>-L20/3 at 1.5 ug/ml in reaction buffer plus 30
µl of microparticles are added and incubated for further 9 min. During incubation an dy
analyte antibody sandwich is formed that is bound to the microparticles. Finally the
microparticles are transferred to the detection chamber of the s system for signal
generation and readout. For calibration 1:2 dilution series of purified CFHR1 (125 ng/ml, 62.5
ng/ml, 31.25 ng/ml, 15.63 ng/ml, 7.81 ng/ml, 3.9 ng/ml, 1.95 ng/ml, 0 ng/ml) are prepared in
MultiAssay Diluent. The equation of the ation curve was calculated by non-linear least-
squares curve-fitting (RCM-Rodbard) and used for converting the signal readout into the
corresponding concentration value. The result is multiplied by the pre-dilution factor to get the
concentration of the respective sample itself.
Cross-reactivity of the applied antibody ch was determined by measuring distinct
concentrations of the potentially cross-reacting serum components CFH, CFHL1, CFHR2,
CFHR3, CFHR4 and CFHR5. For the applied antibody sandwich no cross-reactivity has been
determined for CFHL1, CFHR2, CFHR3, CFHR4, CFHR5 and only minor recognition of CFH
of 4.4 % was determined.
Measurement of serum s from Phase 2 GRI Schizophrenia trial identifies natural
cut-offs within CHFR1
CFHR1 concentrations measured with the CFHR1 ECLIA immunoassay showed a multimodal
distribution which correlates with the on status of the CFHR1 gene.
FIGURE 1 shows the distribution of CFHR1 concentrations in the Phase II serum-samples.
Three groups were identified:
• a group with CFHR1 trations below 8 µg/ml, which correlated with the group
ng a homozygous deletion for CFHR1, and thus does not express CFHR1. The measured
signal is due to the residual cross-reactivity of the CFHR1 ECLIA immunoassay with CFH.
• a group with CFHR1 concentrations above 8 ug/ml and below 28 ug/ml. This group
carries a heterozygous deletion for CFHR1.
• a group with CHFR1 values above 28 ug/ml. This group carries no deletion for CFHR1.
CFHR1 is deleted in about 5-17 % of the population, dependent on ethnic origin [5,9]. In
Caucasians, about 5% of the tion bears a homozygous deletion and about 40% carry a
zygous deletion.
Patient Stratification using CFHR1 natural cut-off
The CFHR1 protein concentrations within each of the three groups shown in FIGURE 1
were analyzed for association with treatment response: In patients carrying the homozygous
deletion, no CFHR1 can be detected; the residual signal is due to cross-reactivity of the assay
with CFH. Within patients carrying the heterozygous deletion of CFHR1 there was no
association of CFHR1 protein concentration with treatment se. Within the patient group
carrying no deletion of CFHR1 there was a trend where a higher CFHR1 concentration
correlated with higher ent response. This tes that the response rate is actually mainly
predicted by the genetic status of CFHR1, with heterozygous or homozygous deletion of CFHR1
translating into to lower response to GRI.
Consequently, for patient stratification, the natural cut-off of CFHR1 ne serum
concentration identified at 28 ug/ml using the ECLIA immunoassay specific for CFHR1
(FIGURE 1) was chosen. ts with CFHR1 serum values below or equal to 28 ug/ml were
stratified as “CFHR1-low”, while patients with CFHR1 serum values above 28 ug/ml were
stratified as “CFHR1-high”. Table 2 shows the distribution of CFHR1-high and CFHR1-low
patients within the different dose groups
CFHR1-high (N=75) and CFHR1-low (N=75) subgroups were analyzed on the primary
endpoint and selected secondary clinical endpoints of the Phase II study and compared to the
non-stratified CFHR1 serum-sample PP subgroup (N=150).
FIGURE 2 shows the analysis on the change in PANSS negative symptom factor score
(PANSS NFS) from baseline over time. Results for the l patient cohort are shown in the
top panel, for the CFHR1-low patients in the middle panel and for the CFHR1-high patients in
the lower panel. For the 10 mg arm the effect size in the CFHR1-high group was -1.01 ed
to -0.42 in the non-stratified group, and for the 30 mg arm the effect size in the CFHR1-high
group was -0.76 compared to -0.47 in the non-stratified group.
FIGURE 3 shows the analysis on the PANSS negative symptom factor score responder
rates, which was a secondary clinical endpoint: Patients with at least a 20% decrease from
ne in the PANSS negative symptom factor score after 8 weeks of treatment were defined as
responders: The response rate for all four treatment arms in the overall serum subgroup is given
as white striped background bars and for the CFHR1-high and CFHR1-low ups as grey
bars. For the 10 mg arm the response rate in the CFHR1-high group was 88% compared to 69%
in the non-stratified group, and for the 30 mg arm the response rate in the CFHR1-high group
was 78% compared to 66% in the non-stratified group.
FIGURE 4 shows the analysis on another secondary clinical endpoint, the clinical global
impression (CGI) of negative symptoms responder rates: For analysis of CGI responder rates,
those patients that were “improved” or “very much improved” were defined as responders: The
response rate for all four treatment arms in the overall serum subgroup is given as white d
background bars and for the CFHR1-high and CFHR1-low subgroups as grey bars. For the 10
mg arm the response rate in the CFHR1-high group was 69% compared to 39% in the nonstratified
group, and for the 30 mg arm the se rate in the CFHR1-high group was 67%
compared to 45% in the non-stratified group.
Antibodies used for the CFH-specific assay
For the development of a specific assay the following monoclonal antibodies were
used: MAB<CFH/CFHR1>M-L20/3 (provider: Thermo ific, cat. no.: GAU 02002)
and MAB<CFH>OX-24 (provider: Thermo ific, cat. no.: MA1-70057).
Ruthenylation of OX24 IgG, stoichiometry 1:3
To a solution of 5 mg/ml monoclonal mouse IgG (clone OX-24, Thermo Scientific, cat. no.:
MA1-70057) 5 mg/ml in 100 mM KPO4, pH 8.5, are added and 125 µg Ruthenium-(bpy)2-
bpyCO-Osu are given to the IgG solution. After 75 min at room ature, ruthenylation is
stopped by addition of 10mM Lysine. For separation of aggregates appropriate fractions of
sample are collected from Superdex 200 size exclusion chromatography.
Purification of CFH from human serum
For use as reference calibrator material, native CFH was purified from human serum by
immunoadsorption using the monoclonal antibody L20/3 specific for CFH and CFHR1
according to following procedure: MAB L20/3<CFH,CFHR1>M-IgG-Spherosil resin is used for
purification. uted (1:4 in 50mM TrisHCl, 150mM NaCl, pH7.5) serum is passed through
Sartoclean CA (0.8µm) and Sartobran P (0.2µm) filter caps. Pretreated serum, loaded on the
column is washed (10mM TrisHCl, 500mM NaCl, 0.05% Tween20, pH 7.5) followed by Q-
sepharose running buffer and eluted in gentle elution buffer (Thermo Scientific) to avoid
degradation of analyte and adsorber. For exclusion of co-bound CFHR1, the eluate from
immunoadsorber is run over a MonoQ column for size dependent n. Pure CFH is applied as
reference calibrator material for the determination of CFH in the s-ECLIA test for serum
CFH values.
Measurement of CFH in human serum or plasma s using the ECLIA immunoassay
ECLIA assay for CFH is developed for the specific measurement of CFH in
human serum or plasma s using the Elecsys® Analyzer.
The following describes the assay procedure for the determination of CFH using the
Elecsys® Analyzer.
The Elecsys CFH immunoassay is an electrochemiluminescence immunoassay (ECLIA)
that functions via the sandwich principle. There are two antibodies included in the assay – a
biotinylated Fab fragment of monoclonal anti-CFH antibody L20/3-Bi (capture dy) and a
ruthenylated monoclonal anti-CFH antibody OXBi Ru (detection antibody) – which form
sandwich assay complexes with CFH in the sample. The complexes are then bound to
solid-phase streptavidin-coated microparticles. The microparticles are magnetically captured
onto the surface of an electrode, and the application of a e to the ode induces
chemiluminescent emission, which is ed by a photomultiplier for readouts. Results are
determined via an instrument-specific calibration curve which is generated by 2-point calibration
and a master curve provided via the reagent barcodecurve. The total time required to perform the
assay is 18 minutes.
Samples are automatically diluted 1:400 in MultiAssay Diluent for s (Roche
03609987-190). Following the applied assay protocol 2, 80 µl of biotinylated <CFH,CFHR1>-
L20/3 at 1.5 µg/ml and 80 µl of ruthenylated <CFH>-OX24 at 1.5 µg/ml both in on buffer
(Hepes50mM, NaCl 150mM; Thesit/Polidocanol 0.1%; EDTA 1mM; bovine serum albumin
0.5%) are incubated with 10 µl of sample. During incubation an antibody analyte antibody
sandwich is formed that is bound to the microparticles. Finally the microparticles are transferred
to the detection chamber of the Elecsys system for signal generation and readout. For calibration
1:4 dilution series of purified CFH (1.25 µg/ml, 312.5 ng/ml, 78.1 ng/ml, 19.5 ng/ml, 0 ng/ml)
(500 µg/ml, 125 µg/ml, 31.25 µg/ml, 7.81 µg/ml, 0 µg/ml)- are prepared in ssay Diluent.
The equation of the calibration curve was is calculated by near least-squares curve-fitting
(RCM-Rodbard) and used for converting the signal readout into the corresponding concentration
value. The result is multiplied by the pre-dilution factor to get the tration of the respective
sample itself.
Cross-reactivity of the applied antibody sandwich was determined by measuring distinct
concentrations of the potentially cross-reacting serum components. For the applied antibody
sandwich cross-reactivity of 2.9% has been determined for CFHR1
Measurement of CFHR1/CFH ratio measured in serum s from Phase 2 GRI
phrenia trial confirms natural cut-offs
CFHR1 concentration and CFH concentration are measured independently with the
CFHR1 and CFH ECLIA immunoassay and allowed calculation of the CFHR1/CFH ratio.
Similar to CFHR1 concentration alone as shown in FIGURE 1, the CFHR1/CFH ratio showed a
multi-modal distribution which ates with the on status of the CFHR1 gene.
FIGURE 5 shows the distribution of the CFHR1/CFH ratio in the Phase II serum-samples.
Three groups were identified:
• a group with a CFHR1/CFH ratio below 0.01, which correlated with the group carrying a
homozygous deletion for CFHR1, and thus does not express CFHR1. The residual signal
measured is due to the low cross-reactivity of the CFHR1 ECLIA immunoassay with CFH.
• a group with a CFHR1/CFH ratio above 0.01 and below 0.08. This group carries a
zygous deletion for CFHR1.
• a group with a CHFR1/CFH ratio above 0.08. This group carries no deletion for CFHR1.
Patient Stratification using the CFHR1/CFH ratio
The CFHR1/CFH ratio within each of the three groups as shown in FIGURE 5 were
analyzed for association with treatment response. For this purpose a CFHR1/CFH ratio below
0.01 was called low (“L”), a CFHR1/CFH ratio above 0.01 and above 0.08 was called medium
(“M”) and a CFHR1/CFH ratio above 0.08 was called high (“H”).
FIGURE 6 shows the analysis on the clinical global impression (CGI) of negative
symptoms der rates: For analysis of CGI responder rates, those patients that were
“improved” or “very much improved” were defined as responders: The response rate for all four
treatment arms in the l serum subgroup is given as white striped background bars and for
the CFHR1/CFH ratio subgroups L, M and H as grey bars. For the L group the response rate was
0% for the Placebo arm and for the 10 mg and 30 mg arm, while in the 60 mg arm the response
was 100%. The L group consisted of a small sample size and thus results need to be interpreted
with some caution. For the M subgroup the response rate with 24% in the Placebo arm and with
% in the 60 mg arm were comparable to the non-stratified group (shaded bar), being 20% for
Placebo and 34% for 60 mg arm. However, in the 10 mg and 30 mg arms the M up
showed lower response rates, 19% (for 10mg) and 28% (for 30mg) ed to that of the
tified group with 39% (for 10mg) and 45% (for 30 mg). The H subgroup showed a
response rate of 20% for the placebo arm and 33% for the 60 mg arm which is comparable to the
response rate of the non-stratified group, being 20% (for placebo) and 34% (for 60 mg). In both
treatment arms, the 10mg arm and the 30 mg arm, the H subgroup showed increased treatment
response rates: a response rate of 69% in the H subgroup compared to 39% in the non-stratified
group for the 10 mg arm, and a response rate of 67% in the H up compared to 45% in the
non-stratified group for the 30 mg arm.
Preparation of DNA samples
DNA s were collected from patients that had given written consent to the collection
and use of DNA samples for exploratory biomarker analysis (170 of 320 ts). DNA samples
were collected as whole blood, collected in a 9 ml EDTA tubes and stored at -20°C. DNA
samples were double coded and de-anonymized. From a sample of 50-200 uL whole blood,
c DNA was extracted using the Roche MagNA Pure 96 LC DNA Isolation System. The
principle of DNA isolation is based on magnetic bead technology. Briefly, the samples are first
lysed by incubation with a buffer containing chaotropic salts and Proteinase K. Magnetic Glass
Particles (MGP) are then added and the DNA binds to their surfaces. Unbound substances are
removed by several washing steps and the purified DNA is eluted. The resultant DNA was
normalized to a concentration of 5 ng/ul using spectrophotometric quantification.
Assay for determination of CFHR1 copy number
A quantitative real-time PCR assay was set up to determine the copy number at the
genomic location at chromosome 1:196796257-196796381 (coordinates according to according
to Genome Build 37, ly GRCh37). Oligonucleotide sequences are given in Seq. Id. No.
8, Seq. Id. No. 9 and Seq. Id. No. 10. The fluorescent probe was modified at the 5’ us with
5’-Fluorescein and at the 3’ terminus with the 3’ al BlackHoleTM Dark Quencher-1.
This CFHR1 copy number assay was set in reference to assays labeled with a separable
fluorescent reporter that detect a sequence known to exist in two copies in a diploid . As
reference assays the TaqMan Copy Number Reference Assays for RNase P H1 RNA and hTERT
genes (Life Technologies, Carlsbad, California), respectively were employed. Results were
considered valid if the resultant CFHR1 copy number from independent duplex setups using
either RNaseP and hTERT reference assays matched.
The PCR reaction contained PCR oligonucleotides at a final concentration of 0.9 uM and
the probe ucleotide at a final concentration of 0.25 uM Real-time PCR reactions were
conducted using a LightCyclerR 480 II real-time PCR system and the following cycling
parameters: 10 min 95° C, 40x [15 sec 95° C, 1 min 60° C], 1 min 40 °C. Employing“Absolute
Quantitation/2nd Derivative Max” instrument settings, the threshold cycle (CT) values were
calculated. Using comparative ve quantitation analysis ∆∆CT [10] the copy number of the
CFHR1 target ce was calculated.
As an external reference for the copy number of CFHR1, DNA from the cell line NA07000
(Coriell Institute for l ch, Camden, New Jersey) was used, which is described to
harbor two copies at a location within the CFHR3-1 deletion [11]. The assay was tested on
multiple cell lines that are described to harbor one or zero copies at the CFHR3-CFHR1 locus
and the specificity of the assay was confirmed.
Assay for determination of rs7542235
DNA samples were genotypes using the Human 1M-Duo Beadchip (v.3) from Illumina
according to the manufactures procedure. Samples were then analyzed for SNP rs 7542235.
Correlation of CFHR1 copy s to Single Nucleotide Polymorphism rs7542235
The Single Nucleotide Polymorphism (SNP) rs7542235, located at genomic position Chr
1: 613 (Genome Build 37, Assembly GRCh37) has been described as a proxy for the
common CFHR1–CFHR3 deletion [12]. This association was shown to be very high for
individuals with european ancestry ethnicity (r2 = 1.00) but less tight for other ethnic groups [5,
TABLE 3 shows the correlation of the customized copy number ion assay with the
SNP rs7542235 allele: consistent with the literature the major A/A allele tags two CFHR1 copies
ed CFHR1 +/+), the allele A/G tags one CFHR1 copy (denoted CFHR1 +/-) and the minor
allele G/G tags zero CFHR1 copies (denoted CFHR1 -/-).
Noteworthy, both samples with CFHR1 -/- and rs7542235 A/A allele are of black, nonhispanic
ethnicity and four out of five patients with CFHR1 +/- and rs7542235 A/A allele are of
non-European ethnicity.
Correlation of CFHR1 copy numbers with CFHR1 protein concentration
FIGURE 7 shows the box plot of the genetic CFHR1 status versus protein concentration
measured using the RBM assay (so mainly detecting CFHR1): CFHR1 +/+ genetic status
correlates with high CFHR1 serum concentration, CFHR1 +/- genetic status correlates with
lower CFHR1 serum concentration and CFHR1 -/- genetic status correlates with the lowest
protein signals (caused by crossreactivity of the assay with CFH).
Correlation of SNP rs 7542235 to CFHR1 n concentration
FIGURE 8 shows the box plot of the allelic status of the SNP rs 7542235 versus protein
concentration measured using the RBM assay (so mainly detecting CFHR1): rs7542235 allele
A/A ates with high CFHR1 serum concentration, 235 allele A/G correlates with
lower CFHR1 serum concentration and rs7542235 allele A/A correlates with the lowest protein
signals (caused by crossreactivity of the assay with CFH).
Patient fication using CFHR1 genetic analysis
FIGURE 9 shows the analysis on the clinical global impression (CGI) of negative
symptoms der rates: For analysis of CGI responder rates, those patients that were
“improved” or “very much ed” were defined as responders: The response rate for those
patients who had consented for a DNA sample is given as white striped background bars and for
the CFHR1-genetic status as grey bars. For the 10 mg arm the se rate in the CFHR1 +/+
group was 57% compared to 39% in the non-stratified group, and for the 30 mg arm the response
rate in the CFHR1 +/+ group was 67% ed to 46% in the non-stratified group.
Description of the Figures
Figure 1: Histogram of CFHR1 concentrations in the Phase II serum baseline samples. The
three curves show the probability density estimates from a clustering algorithm (Mixture of
Gaussians). The clustering analysis assumed 3 different concentration groups based on the
genetic status of CFHR1 expression. 2 natural cut-offs can be identified using the CFHR1
ECLIA immuno assay: at 8 µg/ml and at 28 µg/ml. The 28 µg/ml cut-off (marked with an arrow)
tes the ts with CFHR1 heterozygous or homozygous deletion from those carrying no
deletion and was used for t stratification.
Figure 2: Change of PANSS negative symptom factor score from baseline in all patients
(top panel) and in low patients (middle panel) and CFHR1-high patients (lower panel)
using natural cutoff for CFHR1. Patients with serum tration ≤28 ug/ml were fied as
CFHR1-low and ts with serum concentration >28 ug/ml were classified as CFHR1-high.
Estimates of expected se and standard error bars from MMRM analysis. Dash-dot line and
circle Placebo; solid line and squares 10 mg GRI; solid line and upward-pointing triangles 30 mg
GRI; dashed line and diamonds 60 mg GRI.
Figure 3: Response rates of dichotomized PANSS negative symptom factor score
(response d as ≥20% drop from baseline) at week 8. Shaded background bars are se
rates in overall per protocol population. Solid grey bars are se rates in CFHR1 stratified
subpopulations using natural cut-off for CFHR1.
Figure 4: Response rates of omized CGI-I negative symptoms rating score (response
defined as “much improved” or “very much improved”) at week 8. Shaded background bars are
response rates in overall per protocol population. Solid grey bars are response rates in CFHR1
stratified subpopulations using natural cut-off for CFHR1.
Figure 5: Histogram of the CFHR1/CFH ratio distribution in the Phase II serum baseline
samples. The three curves show the ility density estimates from a clustering algorithm
(Mixture of Gaussians). The clustering analysis assumed 3 different concentration groups based
on the genetic status of CFHR1 expression. 2 natural cut-offs can be identified using the
CFHR1/CFH ratio (measured by the ECLIA assays): 0.01 and 0.08. The samples with a
CFH ratio below 0.01 are indicative of CFHR1 homozygous deletion, samples with a
CFHR1/CFH ratio above 0.01 and below 0.08 are indicative of CFHR1 heterozygous deletion,
and samples with a CFHR1/CFH ratio above 0.08 are indicative of no CFHR1 deletion
Figure 6: Response rates of dichotomized CGI-I negative ms rating score (response
defined as “much improved” or “very much improved”) at week 8. Shaded background bars are
response rates in overall per protocol population. Solid grey bars are response rates in CFHR1
stratified subpopulations using CFHR1/CFH ratio, with L (low) ponding to CFHR1/CFH
ratio below 0.01, M (medium) corresponding to CFH ratio above 0.01 and below 0.08
and with H (high) ponding to CFHR1/CFH ratio above 0.08.
Figure 7: Genetic status for CFHR1 gene versus CFHR1 protein concentration using the
RBM assay (recognizing CFHR1:CFH in a ration 10:1)
Figure 8: Allelic status for SNP rs7542235 versus CFHR1 protein concentration using the
RBM assay (recognizing CFHR1:CFH in a ration 10:1)
Figure 9: Response rates of dichotomized CGI-I negative symptoms rating score (response
defined as “much improved” or “very much improved”) at week 8. Shaded background bars are
response rates in overall per protocol population from those who had consented for DNA sample.
Solid grey bars are se rates in CFHR1 stratified subpopulations using genetic status of
CFHR1 allele.
Description of the Sequences (SEQ IDs):
Seq. Id. No. 1: shows the n sequence of human complement factor H.
Seq. Id. No. 2: shows the protein sequence of human complement factor H related protein
CFHR1.
Seq. Id. No. 3: shows the protein sequence of human complement factor H related protein
CFHR2.
Seq. Id. No. 4: shows the protein sequence of human complement factor H related protein
CFHR3.
Seq. Id. No. 5: shows the protein sequence of human complement factor H related protein
CFHR4A.
Seq. Id. No. 6: shows the n sequence of human complement factor H related protein
CFHR4B
Seq. Id. No. 7: shows the protein sequence of human ment factor H related protein
CFHR5.
Seq. Id. No. 8 shows the oligonucleotide sequence for the d primer used in the customized
CFHR1 copy number assay
Seq. Id. No. 9 shows the oligonucleotide sequence for the reverse primer used in the customized
CFHR1 copy number assay
Seq. Id. No. 10 shows the oligonucleotide sequence for the probe used in the customized CFHR1
copy number assay
Table 1: Highest ranking ker candidates for predicting PANSS and CGI-I negative
symptom scores at week 8 in patients treated with 10 or 30 mg GRI
PANSS CGI-I
Standar-dized Un- Adjusted Un-adjusted Adjusted
Slope adjusted P AUC P P
“CFH” -1.33 0.004 0.17 0.75 0.0003 0.006
CCL23 0.54 > 0.2 > 0.2 0.70 0.004 0.11
(MPIF-1)
FAS 0.45 > 0.2 > 0.2 0.69 0.005 > 0.2
FABP 0.55 0.18 > 0.2 0.68 0.009 > 0.2
Leptin -1.30 0.03 > 0.2 0.63 0.06 > 0.2
CXCL9 0.51 > 0.2 > 0.2 0.67 0.01 > 0.2
MMP-3 1.06 0.03 > 0.2 0.59 0.19 > 0.2
Apolipo- -0.91 0.03 > 0.2 0.58 > 0.2 > 0.2
protein
IL-11 -1.11 0.03 > 0.2 0.46 > 0.2 > 0.2
Table 2: Distribution of CFHR1–high and CFHR1-low patients in different dose
arms using natural CFHR1 cut-off.
Overall CFHR1-low CFHR1- high
N N (%) N (%)
All dose arms 150 75 (50%) 75 (50%)
Placebo 41 21 (51%) 20 (51%)
mg 36 20 (56%) 16 (44%)
mg 38 20 (53%) 18 (47%)
60 mg 35 14 (40%) 21 (60%)
CFHR1 low are patients with baseline CFHR1 serum concentration ≤28 µg/ml,
CFHR1 high group are patients with baseline CFHR1 serum tration >28 µg/ml.
CFHR1 was ined using the ECLIA CFHR1 assay
Table 3: Correlation of SNP 235 alleles to CFHR1 genetic status determined for
DNA samples from the Phase 2 GRI Schizophrenia trial
rs7542235 A/A rs7542235 A/G rs7542235 G/G
CFHR1 -/- 2 0 15
CFHR1 +/- 5 52 0
CFHR1 +/+ 88 0 0
References
Walport, M.J. N. Engl. J. Med., 344, 1058-1066
[2] Isohanni, Eur. Arch. Psychiatry Clin Neurosci., 2000, 250(6), 311-9
Marenco, S, Dev. Psychopathol. 2000, 12(3), 501-27
Jozsi, M and Zipfel, PF Trend in Immunology 2008, 29(8), 380-387
Zhao J. et al, PLoS Genetics, May 2011, Vol. 7(5), e1002079,
Hey, T., et al., Trends Biotechnol. 23 (2005) 514-522;
Tietz entals of clinical chemistry, C.A. Burtis and E.R. d, 5th edition, 2000,
pages 177-194, Principles of Immunochemical Techniques;
[8] Marder SR, et al., J Clin Psychiatry. 1997;58:538–546.
Hagemann et al, Ann Med. 2006, 38 (8),592-604;
Livak & Schmittgen, Methods. 2001 Dec;25(4):402-8.);
Conrad, DF et al., Nature, 2010, 464(7289):704-12.
Raychaudhuri et al. (2011), Nat. Genet. 43, 1232);
Claims (1)
- Claims 1. An in vitro method of predicting a se for patients, having negative or positive symptoms of schizophrenia, if treated with a glycine ke inhibitor (GRI), comprising the 5 steps: i) determining the protein concentration of complement factor H related protein 1 (CFHR1) in a sample of a patient, ii) comparing the protein concentration determined in step i) to a value representative of the protein concentration of complement factor H related protein 1 (CFHR1) in patients, having 10 ve or positive ms of schizophrenia, iii) wherein a higher protein concentration of complement factor H related protein 1 (CFHR1) in the sample of the patient having negative or ve symptoms of schizophrenia is indicative for a t who will derive clinical benefit from this treatment and iv) selecting this treatment for ts having negative or positive symptoms of schizophrenia. 15 2. An in vitro method according to claim 1, wherein the protein concentration of ment factor H related protein 1 ) is determined by ELISA based technology. 3. An in vitro method of predicting a response for patients, having positive or negative symptoms of schizophrenia according to claim 1, wherein the protein concentration of complement factor H related protein 1 (CFHR1) is determined by measuring genetic variants of 20 complement factor H related protein 1 (CFHR1). 4. An in vitro method of predicting a response for patients, having positive or negative ms of schizophrenia, if treated with a glycine reuptake inhibitor (GRI) according to claim 3, wherein the protein concentration of complement factor H related protein 1 (CFHR1) is determined by measuring of genetic variants of CFHR1, either via measurement of copy number 25 ions of CFHR1 or by measurement of a SNP as a proxy for the deletion. 5. The use of an antibody specifically binding to ment factor H related protein 1 (CFHR1) in a method according to any one of claims 1 to 4. 6. An in vitro method according to any one of claims 1 - 4, where the t is affected with schizoaffective disorder. 7. An in vitro method according to any one of claims 1 to 4 and 6, wherein the e reuptake inhibitor is [4-(3-fluorotrifluormethyl-pyridinyl)-piperazinyl]-[5- methanesulfonyl[[(2S)-1,1,1-trifluoropropanyl]oxy]phenyl]methanone. 8. The use of complement factor H related protein 1 (CFHR1) protein as a predictive 5 marker for clinical benefit for patients who are treated with a e ke inhibitor. 9. A method according to any one of claims 1 to 4, 6 and 7 substantially as herein described with reference to any example thereof. 10. A use according to claim 5 or 8 substantially as herein described with reference to any example thereof. FIGURE 1 Homozygous Heterozygous deIetion deletion No n g‘ co 8 o I... E 'llirlllll; ”L" V I : II I I IIIHn-i...'i' 0 10 20 30 40 50 60 CFHR1 [ug/ml] SU BSTITUTE SHEET (RULE 26) FIGURE 2A dPANSSNSFS BASELINE WEEK1 WEEK2 WEEK4 WEEKS WEEKS FIGURE 2B dPANSS.NSFS BASELINE WEEK1 WEEK2 WEEK4 WEEKS WEEKS SUBSTITUTE SHEET (RULE 26) WO 30032 FIGURE 2C dPANSS.NSFS is .3 o BASELINE WEEK1 WEEK2 WEEK4 WEEK6 WEEKS SUBSTITUTE SHEET (RULE 26) 2012/066216 FIGURE3 Oveiali Response 44% 69% 66% 43% 48% 40% 55% 88% 55% 78% 57% 33% Subgroup Response (1021) (8/20) (11l20)(14116) (”mum/13) (8/14) (7/21) 8 . “' I F m . as 8 o: f I g I I g 3 . g a: I o i N ‘ i i l CFHR1 Low High Low High Low High Low High Placebo 10 mg 30 mg 60 mg SU BSTITUTE SHEET (RULE 26) FIGURE4 Overall Response 20% 39% 45% 34% 19% 20% 15% 69% 25% 67% 36% 33% up Response (4/21) (4120) (3f20) (11/16) (5/20) (12/18) (5114) (7(21) 8 -* o 3 . y, ‘ W Wf ” mulllh‘l.‘f'fl'r‘fl 'lfll in il CFHR1 Low High Low High Low High Low High Placebo 10 mg 30 mg 60 mg FIGURE 5 8 L0 0) Y" w 9 0.05 0.10 0.15 0.20 CFHRl/CFH SUBSTITUTE SHEET (RULE 26) FIGURE 6 l Response 20% 39% 45% 34% 0% 24% 20% ms 19% 69% 0% 28% 67% 100% 25% 33% Subgroup Response (014) (4217x490) (W4)(3f16fl1r16) ((1/2) (5i18112l18) (2121(3112x7l21) 93 8 a S o . . CFHR‘l/CFH L M H L M H L M H Piacebo 10 mg 30 mg SUBSTITUTE SHEET (RULE 26) 2012/066216 FIGURE 7 7000 ug/ml concentration 5000 3000 Protein 1000 CFHR1 -/- CFHR1 +/'- CFHR1 +l+ SUBSTITUTE SHEET (RULE 26) FIGURE 8 7000 Lug/mi concentration 5000 3000 1000 rs?542235 NA NG GIG SUBSTITUTE SHEET (RULE 26) FIGURE 9 Overall Response 19% 39% 46% 43% Subgroup response 0% 17% 27% 0% 31% 57% 0% 38% 67% 100% 27% 47% (0/4)(2/12)(4/15) (0/4) (4/13)(8l14) (0/3) (5/13) (8/12) (2/2) (3I11)(7I15) 33 O o S: CFHR1 s SUBSTITUTE SHEET (RULE 26) 27353 Sequence Tisting of Foreign text Sequence Tist Seq. Id. No. 1 protein sequence of human compTement factor H. 1 MRLLAKIICL MLWAICVAED CNELPPRRNT EILTGSWSDQ TYPEGTQAIY 51 KCRPGYRSLG RKGE WVALNPLRKC QKRPCGHPGD TPFGTFTLTG 101 GNVFEYGVKA VYTCNEGYQL LGEINYRECD TDGWTNDIPI CEVVKCLPVT 151 APENGKIVSS AMEPDREYHF GQAVRFVCNS GYKIEGDEEM HCSDDGFWSK 201 EKPKCVEISC KSPDVINGSP ISQKIIYKEN CNMG YEYSERGDAV 251 CTESGWRPLP SCEEKSCDNP YIPNGDYSPL RIKHRTGDEI TYQCRNGFYP 301 ATRGNTAKCT STGWIPAPRC TLKPCDYPDI KHGGLYHENM RRPYFPVAVG 351 KYYSYYCDEH FETPSGSYWD HIHCTQDGWS PAVPCLRKCY FPYLENGYNQ 401 NYGRKFVQGK SIDVACHPGY ALPKAQTTVT CMENGWSPTP RCIRVKTCSK 451 SSIDIENGFI SESQYTYALK EKAKYQCKLG YVTADGETSG SITCGKDGWS 501 AQPTCIKSCD IPVFMNARTK NDFTWFKLND HDGY ESNTGSTTGS 551 IVCGYNGWSD LPICYERECE HLVP DRKKDQYKVG EVLKFSCKPG 601 FTIVGPNSVQ CYHFGLSPDL PICKEQVQSC GPPPELLNGN VKEKTKEEYG 651 HSEVVEYYCN PRFLMKGPNK IQCVDGEWTT LPVCIVEEST CGDIPELEHG 701 PPYY YGDSVEFNCS ESFTMIGHRS ITCIHGVWTQ LPQCVAIDKL 751 KKCKSSNLII LEEHLKNKKE FDHNSNIRYR WIHT VCINGRWDPE 801 VNCSMAQIQL CPPPPQIPNS HNMTTTLNYR DGEKVSVLCQ ENYLIQEGEE 851 ITCKDGRWQS IPLCVEKIPC SQPPQIEHGT INSSRSSQES YAHGTKLSYT 901 ISEE NETTCYMGKW SSPPQCEGLP CKSPPEISHG VVAHMSDSYQ 951 YGEEVTYKCF GPAI KWSH PPSCIKTDCL SLPSFENAIP 1001 VYKA GEQVTYTCAT YYKMDGASNV TCINSRWTGR PTCRDTSCVN 1051 PPTVQNAYIV SRQMSKYPSG ERVRYQCRSP YEMFGDEEVM CLNGNWTEPP 1101 GKCG PPPPIDNGDI TSFPLSVYAP QCQN LYQLEGNKRI 1151 TCRNGQWSEP PKCLHPCVIS REIMENYNIA LRWTAKQKLY SRTGESVEFV 1201 CKRGYRLSSR SHTLRTTCWD GKLEYPTCAK R with variants VAR SEQ 446~449 KTCS —> SFTL (in isoform 2). VAR SEQ 31 Missing (in isoform 2). VARIANT 62 V —> I VARIANT 78 R —> G VARIANT 127 R ~> L T 224 missing Page 1 27353 Sequence Tisting of Foreign text VARIANT 325 C ~> Y VARIANT 400 Q —> K T 402 Y -> H VARIANT 431 C ~> U1 VARIANT 493 T -> 73 VARIANT 536 C ~> 73 VARIANT 551 I -> —{ VARIANT 567 W IV O VARIANT 609 H VARIANT 630 I V E T 673 U) VARIANT 673 nnn< -< VARIANT 850 m IV 7< VARIANT 890 LA lV H VARIANT 893 I l V 73 VARIANT 915 n | V U? VARIANT 936 m l V U VARIANT 950 )O I T 951 -< IV I VARIANT 956 —1 3 VARIANT 959 ('3 -< VARIANT 978 E n VARIANT 997 2 lV —l VARIANT 1007 < VARIANT 1007 < -> L VARIANT 1010 )> ~> T VARIANT 1017 -l -> I VARIANT 1021 -< —> F VARIANT 1043 ('1 —> R VARIANT 1050 N -> Y VARIANT 1059 I ~> T VARIANT 1076 Q —> E VARIANT 1078 R ~> S VARIANT 1119 D —> G VARIANT 1134 V -> G Page 2 27353 Sequence g of Foreign text VARIANT 1142 Y ~> D VARIANT 1143 E VARIANT 1157 ED R VARIANT 1163 (‘3 W VARIANT 1169 VARIANT 1183 E I v n VARIANT 1183 E |v r VARIANT 1183 E w VARIANT 1184 4 |v x VARIANT 1189 r m VARIANT 1191 m IV r VARIANT 1194 m U VARIANT 1197 < > VARIANT 1198 m Iv > VARIANT 1199 m VARIANT 1210 n VARIANT 1215 R —> G VARIANT 1215 R —> Q VARIANT 1225- 1231 YPTCAKR —> FQS VARIANT 1226 P —> S Seq. Id. No. 2 protein sequence of human compTement factor H reIated protein CFHRl. 1 MWLLVSVILI SRISSVGGEA TFCDFPKINH GILYDEEKYK PFSQVPTGEV 51 FYYSCEYNFV SPSKSFWTRI WSPT CFFP FVENGHSESS 101 GQTHLEGDTV QIICNTGYRL QNNENNISCV ERGWSTPPKC RSTDTSCVNP 151 PTVQNAHILS RQMSKYPSGE RVRYECRSPY EMFGDEEVMC LNGNWTEPPQ 201 CKDSTGKCGP PPPIDNGDIT SFPLSVYAPA SSVEYQCQNL YQLEGNKRIT 251 CRNGQWSEPP KCLHPCVISR EIMENYNIAL RWTAKQKLYL RTGESAEFVC 301 KRGYRLSSRS HTLRTTCWDG CAKR with variants VARIANT 157 H VARIANT 159 L T 175 E VARIANT 296 age 3 27353 Sequence 1isting of Foreign text Seq. Id. No. 3 protein sequence of human comp1ement factor H d protein CFHRZ. 1 MWLLVSVILI SRISSVGGEA MFCDFPKINH GILYDEEKYK PFSQVPTGEV 51 FYYSCEYNFV SPSKSFWTRI TCAEEGWSPT PKCLRLCFFP FVENGHSESS 101 GDTV QIICNTGYRL QNNENNISCV ERGWSTPPKC RSTISAEKCG 151 PPPPIDNGDI TSFLLSVYAP GSSVEYQCQN LYQLEGNNQI WSEP 201 CVIS QEIMEKYNIK LKWTNQQKLY SRTGDIVEFV CKSGYHPTKS 251 HSFRAMCQNG KLVYPSCEEK with variant ISOFORM 144—171 ISAEKCGPPPPIDNGDITSFLLSVYAPG -> S (in isoform Short). Seq. Id. No. 4 protein sequence of human comp1ement factor H d protein CFHR3. 1 MLLLINVILT LWVSCANGQV KPCDFPDIKH GGLFHENMRR VGKY 51 YSYYCDEHFE TPSGSYWDYI HCTQNGWSPA VPCLRKCYFP YLENGYNQNY 101 GRKFVQGNST EVACHPGYGL PKAQTTVTCT EKGWSPTPRC IRVRTCSKSD 151 IEIENGFISE SSSIYILNKE IQYKCKPGYA TADGNSSGSI TCLQNGWSAQ 201 PICINSSEKC GPPPPISNGD TTSFLLKVYV YQCQ PYYELQGSNY 251 VTCSNGEWSE PPRCIHPCII TEENMNKNNI KLKGRSDRKY YAKTGDTIEF 301 MCKLGYNANT SILSFQAVCR EGIVEYPRCE with variant VARIANT 71 H —> Y Seq. Id. No. 5 protein sequence of human comp1ement factor H re1ated protein CFHR4A. 1 MLLLINVILT LWVSCANGQE VKPCDFPEIQ HGGLYYKSLR RLYFPAAAGQ 51 SYSYYCDQNF VTPSGSYWDY GWSP TVPCLRTCSK SDVEIENGFI 101 SESSSIYILN EETQYNCKPG YATADGNSSG SITCLQNGWS TQPICIKFCD 151 MPVFENSRAK SNGMWFKLHD TLDYECYDGY ESSYGNTTDS GWSH 201 LPTCYNSSEN CGPPPPISNG QKVY EYQC QSYYELQGSK 251 YVTCSNGDWS EPPRCISMKP CEFPEIQHGH LYYENTRRPY FPVATGQSYS 301 YYCDQNFVTP SGSYWDYIHC TQDGWLPTVP CLRTCSKSDI EIENGFISES 351 SSIYILNKEI GYAT ADGNSSGSIT CLQNGWSAQP ICIKFCDMPV 401 FENSRAKSNG MRFKLHDTLD YECYDGYEIS YGNTTGSIVC GEDGWSHFPT 451 CYNSSEKCGP PPPISNGDTT SFLLKVYVPQ SRVEYQCQSY YELQGSNYVT 501 CSNGEWSEPP RCIHPCIITE ENMNKNNIQL KGKSDIKYYA KTGDTIEFMC Page 4 27353 Sequence Tisting of Foreign text 551 KLGYNANTSV LSFQAVCREG IVEYPRCE Seq. Id. N0. 6 protein sequence of human comp1ement factor H re1ated protein CFHR4B. 1 VILT LWVSCANGQE VKPCDFPEIQ HGGLYYKSLR RLYFPAAAGQ 51 SYSYYCDQNF VTPSGSYWDY IHCTQDGWSP TVPCLRTCSK SDIEIENGFI 101 SESSSIYILN KEIQYKCKPG YATADGNSSG SITCLQNGWS AQPICIKFCD 151 MPVFENSRAK SNGMRFKLHD TLDYECYDGY EISYGNTTGS IVCGEDGWSH 201 FPTCYNSSEK CGPPPPISNG DTTSFLLKVY VPQSRVEYQC QSYYELQGSN 251 YVTCSNGEWS EPPRCIHPCI ITEENMNKNN SDIK YYAKTGDTIE 301 FMCKLGYNAN TSVLSFQAVC REGIVEYPRC E with variant VARIANT 306 G ~> E Seq. Id. No. 7 n sequence of human compTement factor H reTated protein CFHRS. 1 MLLLFSVILI SWVSTVGGEG TLCDFPKIHH GFLYDEEDYN PFSQVPTGEV 51 FYYSCEYNFV SPSKSFWTRI TCTEEGWSPT PKCLRMCSFP FVKNGHSESS 101 GLIHLEGDTV QIICNTGYSL ISCV ERGWSTPPIC SFTKGECHVP 151 ILEANVDAQP KKESYKVGDV LKFSCRKNLI RVGSDSVQCY QFGWSPNFPT 201 SCGP PPQLSNGEVK EIRKEEYGHN EVVEYDCNPN FIINGPKKIQ 251 CVDGEWTTLP TCVEQVKTCG YIPELEYGYV YQHG VSVEVNCRNE 301 YAMIGNNMIT CINGIWTELP MCVATHQLKR CKIAGVNIKT LLKLSGKEFN 351 HNSRIRYRCS HSVC INGKWNPEVD CTEKREQFCP PPPQIPNAQN 401 MTTTVNYQDG EKVAVLCKEN YLLPEAKEIV CKDGRWQSLP RCVESTAYCG 451 PPPSINNGDT TSFPLSVYPP GSTVTYRCQS FYKLQGSVTV TCRNKQWSEP 501 PRCLDPCVVS EENMNKNNIQ LKWRNDGKLY VEFQ CKFPHKAMIS 551 SPPFRAICQE ICE with variants VARIANT 46 VARIANT 216 VARIANT 277 VARIANT 356 T 379 VARIANT 521 VARIANT 529 Page 5 27353 Sequence 1isting of Foreign text seq. Id. No. 8 oiigonucieotide sequence for the forward primer used in the customized CFHRl copy number assay CAG TTC CAA TTG TGT CCA AGT GGA TGT seq. Id. No. 9 oiigonucieotide ce for the reverse primer used in the customized CFHRl copy number assay CAT TTC AAA CAA TCT CCA AGG AGA TGA TG Seq. Id. No. 10 oiigonucieotide sequence for the probe used in the customized CFHRl copy number assay CAA ATG TTC TCA AAG TGT GGT CGC T Page 6
Applications Claiming Priority (5)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP11179058 | 2011-08-26 | ||
EP11179058.0 | 2011-08-26 | ||
EP12164017 | 2012-04-13 | ||
EP12164017.1 | 2012-04-13 | ||
PCT/EP2012/066216 WO2013030032A1 (en) | 2011-08-26 | 2012-08-21 | A method for predicting clinical benefit in the treatment of neurodevelopmental, neurological or neuropsychiatric disorders |
Publications (2)
Publication Number | Publication Date |
---|---|
NZ620075A NZ620075A (en) | 2015-03-27 |
NZ620075B2 true NZ620075B2 (en) | 2015-06-30 |
Family
ID=
Similar Documents
Publication | Publication Date | Title |
---|---|---|
RU2730836C2 (en) | Computational analysis of biological data using a manifold and hyperplane | |
CN109661578B (en) | Protein characterization for differentiating bacterial and viral infections | |
US20210349095A1 (en) | Ros kinase in lung cancer | |
US7597890B2 (en) | Methods and compositions for diagnosing lung cancer | |
WO2010062960A2 (en) | METHODS OF DETERMINING RESPONSIVENESS TO ANTI-TNFα THERAPY IN INFLAMMATORY BOWEL DISEASE | |
WO2019099706A1 (en) | Markers for the diagnosis and treatment of non-alcoholic steatohepatitis (nash) and advanced liver fibrosis | |
CN106133136A (en) | Gout morbidity association molecule, uric acid association disease quality associate evaluation methodology and kits for evaluation, inspection body and the medicine of disease quality with inflammation | |
TW201832780A (en) | Precision treatment of heart failure and cardiorenal syndrome | |
JP2014518624A (en) | Assay reagent for neurogranin diagnostic kit | |
CN104244979A (en) | Methods of treating ankylosing spondylitis using IL-17 antagonists | |
WO2006067056A9 (en) | Compositions and methods for treating mental disorders | |
KR102018369B1 (en) | Mutant Genes as Diagnosis Marker for Amyotrophic Lateral Sclerosis and Diagnosis Method Using the Same | |
US20140065610A1 (en) | Method for predicting clinical benefit in the treatment of neurodevelopmental, neurological or neuropsychiatric disorders | |
Ali et al. | Association of TRPV5, CASR, and CALCR genetic variants with kidney stone disease susceptibility in Egyptians through main effects and gene–gene interactions | |
JP2021170021A (en) | Test method enabling diagnosis specific to early pathology of diabetic nephropathy | |
AU2016273230B2 (en) | Biomarkers for a combination therapy comprising lenvatinib and everolimus | |
NZ620075B2 (en) | A method for predicting clinical benefit in the treatment of neurodevelopmental, neurological or neuropsychiatric disorders | |
US10317401B2 (en) | Methods and compositions for the prediction and treatment of focal segmental glomerulosclerosis | |
WO2015129324A1 (en) | Mental illness marker and use therefor | |
WO2007091328A1 (en) | Method for treatment of lung cancer | |
WO2014207246A1 (en) | New polymorphisms for the diagnosis of idiopathic scoliosis disease | |
KR20220112204A (en) | Method for diagnosing Alzheimer’s disease using brain renin-angiotensin system (RAS) factors | |
CN115315527A (en) | Methods of treatment for subjects with at risk alleles in IL33 | |
WO2015083685A1 (en) | Dlg1/SAP97 GENE SPLICING VARIANT, AND DETECTION OF SCHIZOPHRENIA USING SPLICING VARIANT | |
Wagner et al. | Congress of Clinical Chemistry and Laboratory Medicine |