Likvormarkörer för differentialdiagnostik av demenssjukdomar

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1 Likvormarkörer för differentialdiagnostik av demenssjukdomar Henrik Zetterberg, MD, PhD Docent i neurokemi Avdelningen för psykiatri och neurokemi Göteborgs universitet

2 Likvoranalyser för diagnostik av Alzheimers sjukdom Study design: Follow-up study on MCI (>4 years) MCI n= MCI o AD 56 MCI o MCI 21 MCI o other dementias Cut-off AE42 < 53 pg/ml T-tau > 35 pg/ml Vad har dessa för sjukdomar??? Combination of AE42 and T-tau: Sens. for MCI-AD 95% Spec. non-progr. MCI 83 % +DQVVRQ =HWWHUEHUJ HW DO /DQFHW 1HXURO

3 Överlapp mot andra demensjukdomar T-tau pg/ml 15 1 P-tau pg/ml AD MCI Cont FTD DLB MSA PD PSP AD MCI Cont FTD DLB MSA PD PSP 12 Study design: AD n= 78 MCI-AD n= 15 Cont n= 53 FTD n= 16 DLB n= 15 MSA n= 36 PD n= 46 PSP n= 15 AE42 pg/ml AD MCI Cont FTD DLB MSA PD PSP Sensitivity Specificity T-tau 83 % 89 % P-tau 72 % 95 % AE42 91 % 75 % Comb. 87% 92% (2/3 pos.) PPV NPV Comb. 82% 94% 2OVVRQ HW DO &OLQ &KHP

4 Disease PD versus controls Neurofilament light chain (NFL) Normal NFL in PD (1,11,12,15) Neurofilament heavy chain (NFH) Tau protein (Tau) Beta amyloid 42 (A 42) Glial Fibrillary Acidic Protein (GFAP) Neurofilament Normal NFH in PD (1,15,16) A. Normal Tau in PD (13,58,59,6,61,62). B. Not significantly increased Tau in PD (31) C. Not significantly decreased Tau in PD (1,33) A. Normal A 42 in PD (39,41,42,62). B. Not significantly decreased A 42 in PD (31). Normal GFAP in PD (12,13) MSA, MSA-P, MSA-C versus controls Increased NFL in MSA, MSA-P and in MSA-C (1,12) A. Increased NFH in MSA, MSA-P and in MSA-C (1,15,16); B. Normal NFH in MSA-P and increased NFH in MSA-C but not statistically verified (16). Increased Tau in MSA, MSA-P and MSA-C (1,13,15) A. Decreased A 42 in MSA, Normal GFAP in MSA-P and MSA-C (41); B. MSA, MSA-P and Normal A 42 in MSA, MSA- MSA-C (12,13) P and MSA-C (42). PSP versus controls Increased NFL in PSP (12) Increased NFH in PSP (16) Normal Tau in PSP (34,35) Normal A 42 in PSP (41,42) Normal GFAP in PSP (12) CBD versus controls - Normal NFH in CBD (16) A. Increased Tau in CBD (34, 36) B. Normal Tau in CBD (35) - - MSA-P contra MSA-C No difference in NFL between MSA-P and MSA-C (11,12,15) A. No difference in NFH between MSA-C and MSA-P (15,16); B. Increased NFH in MSA-C compared wih MSA-P but not statistically verified (16). No difference in Tau between MSA-P and MSA-C (13,15) No difference in A 42 between MSA-P and MSA-C (41,42) No difference in GFAP between MSA- P and MSA-C (12,13) PD contra MSA, MSA- P, PSP, CBD Higher NFL in MSA, MSA-P and PSP than in PD (1,11,12) A. Higher NFH in MSA, MSA-P and PSP than in PD and CBD. No difference in NFH between PD and CBD (1,16); B. No difference in NFH between PD and MSA-P but not statistically verified (16). Higher Tau in MSA and MSA-P than in PD (1,12) A. Decreased A 42 in MSA but not in PD and PSP (41); B. No difference in A 42 between PD, MSA and PSP (42). No difference in GFAP between PD, MSA and PSP (12,13) MSA contra PSP No difference in NFL between MSA and PSP (11,12) No difference in NFH between MSA and PSP (16) Higher Tau in MSA than in PSP (indirect conclusion, no study) No difference in A 42 between MSA and PSP (42) No difference in GFAP between MSA and PSP (12) CBD contra PSP - Higher NFH in PSP than in CBD (16) A. Higher Tau in CBD than in PSP (34); B. No difference in Tau between CBD and PSP (35) - - MSA contra CBD - Higher NFH in MSA than in CBD (16) Contradictory results (indirect conclusion, no studies) - - Parkinson s disease (PD), multiple system atrophy (MSA), multiple system atrophy with predominantly parkinsonism (MSA-P), multiple system atrophy with predominantly cerebellar symptom corticobasal degeneration (CBD) Radu Constantinescu, in manuscript

5 Frontallobsdemens T-tau pg/ml 15 1 P-tau pg/ml AE42 pg/ml AD MCI FTD AD MCI FTD AD MCI FTD 2OVVRQ HW DO &OLQ &KHP 2WKHU FDQGLGDWH PDUNHUV VXFK DV 6E DQG QHXURILODPHQW KDYH IDLOHG WR UHDFK KLJK HQRXJK VHQVLWLYLW\ DQG VSHFLILFLW\ UHTXLUHG IRU XVH DV GLDJQRVWLF )7' WHVWV

6 Search for new CSF protein markers using SELDI-TOF CSF is applied to protein chips and protein binding is analyzed by MS Different types of chips bind different types of proteins Hydrophobic Anionic Cationic Metal Ion Normal Phase

7 Search for new CSF protein markers using SELDI-TOF About 2 protein peaks can be identified and analyzed Differences in protein peaks between controls and FTD are quantified Control cont. CM1 SPA FTD FTD CM1 SPA cont. Control Q1 SPA FTD FTD Q1 SPA cont. Control IMAC3 SPA FTD FTD IMAC3 SPA CM1 Q1 IMAC C cont. Control H5 SPA FTD FTD H5 SPA H

8 SEARCH FOR BIOMARKERS FOR FRONTOTEMPORAL DEMENTIA USING SELDI Study design: CSF samples from 16 FTD cases 12 healthy controls Protein chips CM1 weak cation exchange Q1 strong anion exchange IMAC3 immobilized metal affinity capture H5 reversed phase / hydrophobic interaction Matrix CHCA = α-cyano-4-hydroxy cinnamic acid Small proteins < 15 kda SPA = sinapinic acid Large proteins > 1 kda Results: About 2 peaks detected, possible to quantify Primary analysis: 42 peaks with p<.1 Exclusion of peaks with low signal-to-noise ratio differentially charged peaks 1 peaks for PLS-DA analysis

9 SEARCH FOR BIOMARKERS FOR FRONTOTEMPORAL DEMENTIA USING SELDI =HWWHUEHUJ HW DO ([S 1HXURO

10 Examples of individual peaks in FTD and controls 6, M , M 564 5, 4, 3, 3, 2, 2, 1, 1,,, FTD Controls FTD Controls 3, M , M , 2, 2, 1, 1,, FTD Controls, FTD Controls =HWWHUEHUJ HW DO ([S 1HXURO

11 Sex Age T-tau P-tau AE42 M 3689 M 564 M 526 M 6251 M M M 1254 M M 6247 M Quantitative differences between FTD and controls Variable =HWWHUEHUJ HW DO ([S 1HXURO CoeffCS[1](Diagnosis)

12 PLS-DA loadings for all variables Control M M 1254 M 6247 M 526 M AE42 P-tau M 3689 M 564 FTD M 6251 M T-tau M Loadings for: 1 SELDI-TOF peaks 3 CSF markers (T-tau, P-tau Aβ42) FTD-2 C-9 C-3 FTD-12 FTD-11 C-8 FTD-14 FTD-16 FTD-5 FTD-9 FTD-1 FTD-4 FTD-13 FTD-15 FTD-1 FTD-7 FTD-8 FTD-6 C-2 C-5 C-11 C-7 C-6 C-1 C-4 C-1 C-12 FTD =HWWHUEHUJ HW DO ([S 1HXURO

13 Principles for identification of SELDI-TOF peaks AD AD AD AD On-chip tryptic digestion Normal SELDI-TOF Normal Normal #97 RT:11.3 AV:1 NL:5.56E6 F: + c d Full ms @35. [ ] 1 Peptide mapping + PCI1-MS/MS sequencing ProteinChip to monitor purification Relative Abundance m/z SDS-PAGE 21 kda 14 Tryptic digestion Micro-fractionation using matched Biosepra columns ATL-1 Normal 6

14 GRPEAQPPPLSSEHKEPVAGDAVPGPKDGSAPEVRGA b Marker 3689: Neurosecretory protein VGF % b y y y b b y b HH y y FQDKQYSSHHTAE 14.5 Marker 6251: Chromogranin B % HT b QDK b y y 5 b b 5 y 7 b 6 y y a b a b y 1 b m/z

15 FTD versus controls VGF Truncated cystatin C chromogranin B transthyretin S-cysteinylated transthyretin Sensitivitet 94% Specificitet 83% =HWWHUEHUJ HW DO ([S 1HXURO

16 AD mot kontroller och FTD helt oberoende studie p-value p- value M/Z ID AD vs Healthy Level in AD control AD vs FTD Integral membrane protein 2B CT.39 Up n.s Down Up Chromogranin A peptide (Vasostatin II).11 Down VGF peptide.2 Down n.s Down n.s <.1 Down Apo C-1 (2 aa deleted from NT) <.1 Down n.s Down C3a des-arg.4 Up n.s Down n.s Cystatin C -8aa from NT <.1 Up < Cystatin C <.1 Down < Pancreatic ribonuclease <.1 Up n.s <.1 Down n.s. Suboptimala sensitivitets- och specificitetssiffror för att skilja FTD från AD Simonsen et al., Dem Ger Cogn Dis (27) 24:434-44

17 Konklusion 1 9L VDNQDU IRUWIDUDQGH HQ ELRNHPLVN PDUN U VRP NDQ GLIIHUHQWLHUD )7' IUnQ $' Sn HWW EUD VlWW«Det är lätt att skilja ut AD från PD och PSP med standardlikvormarkörerna (T-tau, AE42 och P-tau) MSA och DLB är svårare Det saknas likvormarkörer för att differentiera PD, PSP, MSA och DLB på ett bra sätt

18 Vad är på gång, november 27??? 7DX LVRIRUPP QVWHU Borroni et al., Neurobiol Aging 27, IP-WB Kvoten mellan Tau 33 kda och Tau 55 kda kraftigt sänkt i PSP jämfört med - kontroller - AD - PD - CBD - LBD D6\QXFOHLQ Brit Mollenhauer et al., ELISA (submitted) Och: 7RNXGD HW DO %%5& LQ SUHVV /lww VlQNW DV\QXFOHLQNRQFHQWUDWLRQ L &6) L 3' D6\QXFOHLQDJJUHJDW ELISA från Biosource Funkar ej för CSF 7'3 Virginia Lee och John Trojanowski Den gemensamma länken mellan ALS och FTD?

19 TDP-43 = transactive response (TAR)-DNA-binding protein 43 verkar vara patogenetiskt involverat i både sporadisk ALS och i olika FTD-former. Normalt intranukleärt men vid dessa tillstånd extranukleära inklusionskroppar. FTD (hjärna) ALS (ryggmärg)

20 Konklusion 2 *HQHUHOO SURWHRPLN ± LQJD VWRUD IUDPVWHJ lqqx *RWW RP NDQGLGDWPDUN UHU 7'3 DV\QXFOHLQ 3URJUDQXOLQ WDXNYRWHU PP HU NOLQLVN IRUVNQLQJ EHK YV VlUVNLOW LQVDPOLQJ DY VWRUD RFK YlONDUDNWlULVHUDG SDWLHQWPDWHULDO PHG OLNYRU KHOEORG SODVPD RFK JlUQD KMlUQYlYQDG

21 Tack för uppmärksamheten! Dept of Neurochemistry and Psychiatry Sahlgrenska Academy, Göteborg University Kaj Blennow Erik Portelius Ulf Andreasson Ann Brinkmalm Gunnar Brinkmalm Anders Wallin Clinical Memory Research Unit, Department of Clinical Sciences Malmö, Lund University, Sweden; Lennart Minthon Elisabet Londos Oskar Hansson Chiphergen Anja Hviid Simonsen Vladimir N Podust Excellent technical assistance Åsa Källén, Shirley Fridlund, Mona Thunell, Sara Hullberg, Monica Christiansson Research Support The Swedish Research Council, the Swedish Council for Working Life and Social Research, the Göteborg Medical Society, the Swedish Medical Society, the Swedish Society for Medical Research, Pfizer inc. s grant for clinical research in neuroscience, Stiftelsen Gamla Tjänarinnor, Alzheimerfonden