blank slide/colon data. the data: expresion level of gene i in sample j the data sample 136 1 358...
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Blank slide/colon data
CLUSTER ANALYSIS OF DNA ANDCLUSTER ANALYSIS OF DNA AND ANTIGEN CHIP DATA ANTIGEN CHIP DATA
EYTAN DOMANYEYTAN DOMANY
DEAD SEA, OCT 2002 DEAD SEA, OCT 2002
THE METHOD 1
T (RESOLUTION)
YOUNG OLD
THE METHOD: CLUSTER ANALYSIS.
N OBJECTS: BREAK THEM INTO GROUPS ON THE BASIS OF SIMILARITY:
DENDROGRAM
THE METHOD 2
THE METHOD: CLUSTER ANALYSIS.
N OBJECTS: BREAK THEM INTO GROUPS ON THE BASIS OF SIMILARITY
1. OBJECTS = GENES:
GENES WITH SIMILAR EXPRESSION PROFILES MAY BE CO-REGULATED
PROVIDE GUESS FOR ROLE OF PROTEINS
2. OBJECTS = SAMPLES:
CLASSIFY TUMORS, DIAGNOSIS,
PROGNOSIS, THERAPY
THE PROBLEM RAISED BY ESHEL BEN-JACOB:
CLASSIFYING THE PATIENTS ON THE BASIS OF EXPRESSION OF THOUSANDS OF GENES DOES NOT WORK, SINCE MOST GENES ARE NOT RELEVANT TO THE QUESTION OF INTEREST AND INTRODUCE ONLY NOISE.
THE SOLUTION: WORK WITH SMALL SUBSETS OF GENES AND SAMPLES.
COUPLED TWO-WAY CLUSTERING
Getz et al PNAS (2000)
Califano et al , Proc. Int. Conf. Intell. Syst. Mol. Biol. (2000).Y. Cheng and G. M. Church, Proc. Int. Conf. Intell. Syst. Mol. Biol. (2000)
IDENTIFY (CORRELATED) GROUPS OF GENES AND USE THEIR EXPRESSION LEVELS TO STUDY (CLUSTER) THE SAMPLES.
Astrocytoma(II)Secondary GBM
Primary GlioBlastoMaCell Lines
GE
NE
S
S2S3
T
S1(G1)
G12
G5
Coupled Two-Way Clustering (CTWC)
of 358 Genes and 36 Samples
GLIOBLASTOMA: M. Hegi et al CHUV, G. Getz
glioblastoma
CLONTECH ARRAYS
G1(S1)
AB004904 STAT-induced STAT inhibitor 3
M32977 VEGF
M35410 IGFBP2
X51602 VEGFR1
M96322 gravin
AB004903 STAT-induced STAT inhibitor 2
X52946 PTN
J04111 c-jun
X79067 TIS11B
S11S12
S14
S10
S13S1(G5)
Super-Paramagnetic Clustering of All Samples
Using Stable Gene Cluster G5
Fig. 2B
S1(G5)
AB004904 STAT-induced STAT inhibitor 3 M32977 VEGF
M35410 IGFBP2
X51602 VEGFR1
M96322 gravin
AB004903 STAT-induced STAT inhibitor 2
X52946 PTN
J04111 c-jun
X79067 TIS11B
THE GENES OF G5:
VEGF AND ITS RECEPTORS – INSTRUMENTAL INANGIOGENESIS; INDUCED GROWTH OF BLOODVESSELS, ESSENTIAL FOR GROWTH BEYOND ACRITICAL SIZE. THE COEXPRESSION OF IGFBP2WAS INDEPENDENTLY VERIFIED; 1ST EVIDENCEFOR POSSIBLE ROLE IN ANGIOGENESIS.
THE GENES OF G5
G8
G1(S1)
G3
COLON CANCER: 18 PAIRED CARCINOMA/NORMAL
4 PAIRED ADENOMA/NORMAL
Notterman et al Cancer Res. (2001); Getz et al, Bioinformatics (in print)
colon paired G1(S1)
COLON CANCER: 18 PAIRED CARCINOMA/NORMAL 4 PAIRED ADENOMA/NORMALNotterman et al Cancer Res. (2001)
S1(G8): tumor/normal S1(G3): protocol A /protocol B
COLON CANCER: 18 PAIRED CARCINOMA/NORMAL 4 PAIRED ADENOMA/NORMALNotterman et al Cancer Res. (2001)
S1(G8): tumor/normal distance matrix
BREAST CANCER DATA (BOTSTEIN/BROWN LAB PEROU ET AL, NATURE 2000) I.Kela, G. Getz
20 patients before/after chemotherapy. 10 of the “before” samples are in cluster b; all 3 successful treatments’ samples in this group.
Intermediate expression level of the G46 genes may serve as a marker for a relatively high success rate of the doxorubicin treatment
Predicting response to doxorubicin treatment;successful for 3/20 patients
S1(G46)
survival S1(G33) Sorlie
BREAST CANCER DATA (BOTSTEIN/BROWN LAB),
Sorlie et al, PNAS (2001); Getz et al, Bioinformatics (in print)
Cluster (a): high expression levels of the genes of G33,low survival, mutant p53.
predictor of survival.
S1(G33) survivalp53 status
nointerpret
BREAST CANCER DATA (BOTSTEIN/BROWN LAB),
Sorlie et al, PNAS (2001)
Gene cluster G36 inducesclear partition to two classes of no known clinical interpretation
skin cancer, UV
NORMAL HUMAN EPIDERMAL KERATINOCYTES (NHEK)SQUAMOUS CARCINOMA CELLS (SCC)
IRRADIATE (2m) BY UVB; MEASURE EXPRESSION VS TIMENHEK: UV t =0.5, 3, 6, 12, 24 hours NO UV t = 0, 0.5, 12, 24 SCC: UV t = 0, 6, 12
UV INDUCES DNA DAMAGE, WHICH ELICITS APOPTOTICRESPONSE. NHEK RESIST APOPTOSIS BY SECRETION OFSURVIVAL FACTORS. THIS RESISTANCE TO APOPTOSISMAY PROMOTE EMERGENCE OF MALIGNANCY.
Givol, Rechavi, Dazard,... Hilah Gal
Squamous Carcinoma Cells + UVB (SU)Squamous Carcinoma Cells control (SC)
Normal Keratinocytes control (KC)Normal Keratinocytes + UVB (KU)
Cluster S1(G28)(22 genes)
Reordered Samples
Re
ord
ere
d G
en
es
SC
0h
KC
12h
KC
0h K
C0.
5h
KU
V0.
5h
KC
24h
SU
V6h
SU
V12
hK
UV
3h
KU
V6h
KU
V12
h
KU
V24
h
UV/NON UV SEPARATION INDUCED BY G(28):DNA REPAIR (GADD45A,B); ANTIOXYDANT (MT1G)GROWTH FACTORS, INFLAMMATORY MEDIATORS
S1(G28)
S1(G18) ; TUMOR/NORMAL(5 genes)
Re
ord
ere
d G
en
es
Reordered Samples
KC
12h
KC
0h
SC
0hKC
0.5h
KU
V0
.5h KC
24h
SU
V6h
SU
V1
2hKU
V3h
KU
V6h
KU
V12
h
KU
V24
h
S1(G24) TUMOR/NORMAL(31 genes)
KC
12h
KC
0h
SC
0hK
C0.
5h
KU
V0.
5h
KC
24h
SU
V6h
KU
V3h
K
UV
6h
KU
V1
2h KU
V24
h
Reordered Samples
Re
ord
ere
d G
en
es
SU
V12
h
Squamous Carcinoma Cells + UVB (SU)Squamous Carcinoma Cells control (SC)
Normal Keratinocytes control (KC)Normal Keratinocytes + UVB (KU)
PRO-APOPTOTIC GENES (PARP, CAS)
HIGH IN NHEK, ELEVATED WITH UV
S1(G24), S1(G18)
IRUN COHENFRANCISCO QUINTANAGuy HeadGaddy GetzHila ShtarkDafna TsafrirGadi Elitzur
DIABETES, ARTHRITIS:
Antigen chips
ANTIGEN CHIPS
SAMPLES: N = 40 SERA FROM20 HEALTHY SUBJECTS + 20 DIABETES TYPE 1
78 TESTED ANTIGENS (1 BLANK) EACH WITH TWO “MARKERS”, IgG + IgM AND IgM
MEASURE Aij - REACTIVITY OF SERUM j TO ANTIGEN i
M = 176 MEASUREMENTS PER SAMPLE
THE DATA FORM A 176 X 40 ARRAY
EACH ONE OF THE 40 SUBJECTS IS REPRESENTED BY
176 NUMBERS – THE REACTIVITY OF HIS SERUM WITH
THE 176 ANTIGENS. THESE 176 NUMBERS A1,j , A2,j ,...A176,j
CONSTITUTE THE “REACTIVITY PROFILE” OF SUBJECT j
QUESTION: CAN ONE IDENTIFY PATTERNS OF SIMILARITY BETWEEN THE REACTIVITY PROFILES OF SUBJECTS WITH DIABETES? DO THEY FORM A DISTINCT GROUP FROM HEALTHY SUBJECTS?
ANSWER: NO SEPARATION INTO DIABETES VS HEALTHY IS FOUND WHEN WE USE ALL ANTIGENS TO CHARACTERIZE THE SUBJECTS.
CLUSTERING 176 ANTIGENS, USING THEIR REACTIVITIES WITH 40 SERA:
ANTIGEN CLUSTERS:
RANDOM DATA
THE ANTIGENS FORMDISTINCT GROUPS. THEREIS STRUCTURE IN THEIR REACTIVITY PATTERNS.
USE THE STABLE (SIGNIFICANT) ANTIGEN CLUSTERS,ONE AT A TIME, TO CLUSTER THE SUBJECTS.
USING ANTIGEN CLUSTER 1(Insulin G+M, Collagen1 – both)WE GET A GOOD CLASSIFIER:A “DIABETES CLUSTER”CATCHES 17/20 OF THE DIABETES 4/21 MISTAKES
USING A “MAJORITY VOTE” OF 5 CLASSIFIERS GET 90%
Projects, Collaborators, Students/Postdocs Cancer: Colon* D. Notterman G. Getz*, M.Mashiah*, H. Gal* Breast* D. Botstein I. Kela*, G.Getz* Glioblastoma* M.Hegi, S. Goddard G. Getz* Skin* D. Givol*, G.Rechavi, J. Dazard*H. Gal* Leukemia E. Canaani* O. Ravid*, G.Getz*, H.Agrawal*P53 primary targets* D. Givol*, K.Kannan,G.Rechavi G. Getz*, I.Kela*P73 primary targets* D. Givol*, G. Rechavi I. Kela*MutP53 as oncogen V. Rotter* O. Ravid*Bone development D. Gazit O. Ravid*Antigen Chips: Diabetes* I. Cohen*, F. Quintana* G. Getz*, G. Hed, D. Tsafrir*, Arthritis I. Cohen, F. Quintana* I. Tsafrir*,H.Shtern*,G.Elitzur*Neurotransmitters M. Levite* D. Tsafrir*,I. Tsafrir*Tissue dependence D. Lancet* H. Shtern*Apoptosis, IL6 L.Sachs*,Y.Lotem*,D.Givol* H. Gal*Meiosis in yeast M. Primig M. Katzenelenbogen*Yeast cell cycle* M. Zhang G. Getz*, E. LevineProtein Struct. Classif.* M. Vendruscolo,G.Getz*Low-T phase, SpinGlass* D. Stauffer, P. Young, G. Hed A. Hartmann, M.Palassini SPC* M. Blatt, S. Wiseman H. Agrawal, N. Shental*CTWC* G. Getz*, E. Levine,O.Barad* *re/preprint available *Weizmann Inst. *Currently postdoc/student
collaborators
THE COUPLED TWO WAY CLUSTERING METHOD
SUCCESSFULLY IDENTIFIED RELEVANT STRUCTURE AND
MEANING IN CANCER RELATED GENE MICROARRAY DATA.
CTWC SERVER: http//ctwc.weizmann.ac.il
www.weizmann.ac.il/home/fedomany/
www.weizmann.ac.il/physics/complex/compphys
SUMMARY
Summary
DNA CHIPS PROVIDE A POWERFUL TOOL TO STUDY GENEEXPRESSION; ADVANCED METHODS OF ANALYSIS MAYHAVE FAR REACHING CLINICAL & SCIENTIFIC IMPLICATIONS
FUNDING: ISF, GIF, Ridgefield Found., Levine Found, NIH, EC