Doc618 - Dr Steffan HaelanWien2007neu1

metavectum
Recklinghausen
Modern Moleculare Diagnostic
in Oncology
Aims of Molecular
Oncology
 Early Diagnosis of Tumors
 Detection of disseminated Tumor-Cells and
Micrometastasis
 Deciding of therapeutic Aims und Therapy
Resistance
 Monitoring of Therapy
 Individualisised Therapy
Tumorprogression by
genetic Alterations
Dissemination/Metastasis
Cancer is caused by genetic
Alteration
 cancer cells obtain
Unkontrolled
Growth Signal
unnormal features
Unresponsive against
growth inhibitors
Loss of
Apoptosis
Angiogenesis
Metastasis creation
Unlimited Replication
Genetic Changes
in Tumor-Cells
Genes for Growth Factors
Erb-b2, EGFR, c-Kit, PDGFR, K-ras
Replications facilitating Genes
Telomerase, c-myc
Angiogenesis facilitating Genes
VEGF
Apoptosis facilitating Genes
Bcl-2, p53
Circulating Tumor-cells (CTC)
 Risk-Factor for recurrance and metastasis.
 indicate risk for micro-metastasis.
 Easier indentification of patients profiting from
adjuvant therapy.
Suggested further reading:
1. Cristofanilli M. et al. Circulating tumor cells, disease progression, and survival in metastatic breast cancer. N Engl J Med. 2004 Aug 19;351(8):781-91.
2. Allard WJ et al.: Tumor cells circulate in the peripheral blood of all major carcinomas but not in healthy subjects or patients with nonmalignant diseases
Clin Cancer Res. 2004 Oct 15;10(20):6897-904.
Molekular Charakterisation of
CTCs
 Isolation of mRNA from CTCs
 Measuring different Gene-Expression
 Quantitative real-time PCR of molekular Tumor-Marker (e.g.):
• Cytokeratine
• PSA (Prostata), G250 (kidney), AFP (liver)
• Growth / Cell-cycle-Regulating Gene (c-myc, erbb2,
Telomerase)
Effect of Soy on Gene-Expression
disseminated Tumor-Cells

Five Patients per group suffering from metastastastic Prostate-, Ovaryand Breast -Ca received Chemo + fermented soy

Clinical response in all patients

Question: Can molecular factors predict clinical outcome of CaTherapy by fermented Soy ?

Clincial : Bestimmung der modulierende Wirkung von Soja auf die
Gen-Expression vor und nach 100 Tagen Einnahme des SojaExtraktes
Funktion der Östrogenrezeptoren
(ER) in der Zelle
ER-alpha
a
a
ERE
DNA-Doppelstrang
Zellteilungsaktivität wird gesteigert (Wucherung)
ER-beta
ß
ß
ERE
DNA-Doppelstrang
Zellteilungsaktivität wird verringert
Expression der Östrogenrezeptoren
(ER) alpha und beta
Normale Zellen
Brust / Ovar
Prostata
Tumorzellen
mit Soja-Extrakt
Tumorzellen
unbehandelt
konstantes Verhältnis
Anstieg
Absinken
Anstieg
ß
Absinken
a
ß
ER-beta
ER-alpha
ER-beta
a
ER-alpha
ß
ER-beta
a
ER-alpha
Estrogen-receptor beta is induced
by Haelan 951
Haelan increases ER2 expression
in circulating tumor cells of P-CA patients (n=5)
relative gene expression
600
vor H
nach H
500
400
300
200
100
0
cases
Modulation of estrogen-receptor
expression by Haelan 951
relative gene expression
Circulating Tumor Cells (CTCs) of prostate cancer patients were analysed for
expression of ER-alpha and ER-beta
600
500
400
300
200
100
0
vor H
nach H
ER 1
vor H
nach H
ER 2
-> Expression of ER-beta was significantly induced by Haelan 951
Modulation of estrogen-receptor
expression by Haelan 951
Mamma-Ca. patients
n= 5
all shown
breast cancer patients
are
ER-beta positive +
relative Genexpression
Estrogenreceptor
ER-beta
Estrogenreceptor-beta (ER-beta)
Expression
= No Haelan951
= Haelan951 application
33,9
Healthy persones
without Haelan951
with
Haelan951
6,2
Healthy persones n=7
Mamma-Ca. patients n=5
Relative Genexpression
Mamma-Ca.
Patients
Estrogen-receptor beta is induced
by Haelan 951
Haelan increases ER2 expression
in circulating tumor cells of ovarian cancer patients (n=5)
relative gene expression
100
vor H
nach H
50
0
cases
Modulation of estrogen-receptor
expression by Haelan 951
Circulating Tumor Cells (CTCs) of ovarian cancer patients were analysed for
expression of ER-alpha and ER-beta
Induction of ER1 and ER2 by Haelan
relative gene expression
200
100
0
vor H
nach H
ER 1
vor H
nach H
ER 2
-> Expression of ER-beta was significantly induced by Haelan 951
Stoffwechselweg der Apoptose
Duales Signal von c-myc:
• Der apoptotische Stoffwechselweg wird blockiert durch bcl2 / bax und c-myc fördert Proliferation
• Nach Verlust des bcl2 Überlebenssignals, fördert c-myc die Apoptose
proliferation
mitogens
c- myc
apoptosis
blocking
Bcl2
NF - kB
Bax
(survival signals)
Apoptotic conditioning induced
by Healan 951
CTCs of prostate cancer patients showed an increased gene expression ratio of
apoptosis-triggering genes bax/bcl2 after Haelan951-therapy
Healan induces pro-apoptotic condition
in circulating tumor cells of P-CA patients
vor H
nach H
bax/bcl2 ratio
70
60
50
10
5
0
cases
Apoptotic conditioning induced
by Healan 951
Mamma-Ca. patients
without Haelan951
n= 5
Haelan951 application 100 days
relative Genexpression
BAX/BCL2
no significant
change
1,2
1,1
Apoptotic conditioning induced
by Healan 951
Ovarian-Ca. patients n=6
relative gene expression
10
5
0
vor H
Bax/Bcl2
nach H
Bax/Bcl2
Apoptotic conditioning induced
by Healan 951
Haelan increases BAX expression in circulating
tumor cells of ovarian cancer patients (n=5)
relative gene expression
30
vor H
nach H
20
10
0
cases
The p53 Signaling Pathway
Bax
block
MDM2
p53
Bcl-2
Caspases
block
Cell cycle inhibitor
Cyclin
D1
CDK4
Cyclin E
CDK2
Apoptosis
p21
(Ovarian Ca.)
block
Cell Cycle
Progression
p21 Involvement in Cell Cycle
p53
p21 CIP1
Cell cycle inhibitor
Mitosis
block
Cyclin A/B
CDK1
CDK2
M-phase
entry
G1
DNA-Synthesis
M
G2
S
Chromosomes
Duplication
Cellcycle Inhibitor P21 is induced
by Haelan951
Prostata-Ca. patients n=5
Haelan951 (100) Days)
without Haelan951
56,3
relative Genexpression
37,8
12,4
7,1
1,9
0,5
0,9
cases
1,0
0,6
0,9
Cellcycle Inhibitor P21 is induced
by Haelan951
without Haelan951
Haelan951 (100 days)
Mamma-Ca. patients n=5
332,5
211,9
7,36
4,5
0,11
1,11
cases
2,28
scale unit progressive
15,0
14,3
relative Genexpression
15,8
Cellcycle Inhibitor P21 is induced
by Haelan951
Mamma-Ca. patients
without Haelan951
n= 5
Haelan951 application 100 days
relative Genexpression
p21 CIP
increased
2,0
13,1
Cellcycle Inhibitor P21 is induced
by Haelan951
Haelan increases P21expression in circulating tumor cells
of ovarian cancer patients (n=5)
relative gene expression
150
vor H
nach H
100
50
0
cases
Schlussfolgerungen
• Alle Patienten zeigten auf die Gabe fermentierten Soja-Extraktes
klinischen Response
• Fermentiertes Soja-Extrakt fördert den pro-apoptotischen Zustand der
Prostata-Karzinomzellen durch Herunterregulation von Bcl2. Dies dürfte
die Zellen sensitiver machen für eine Chemotherapie und somit für eine
Kombinationstherapie mit konventionellen Chemotherapeutika sprechen.
• Zugabe von fermentiertem Soja induziert die Expression des
antiproliferativen ER-beta bei Prostata- und Ovar-Ca Patienten.
• Fermentiertes Soja-Extrakt induziert die Transkription des p21, welches
zu einem Stillstand des Zellzyklus führt und damit das Krebswachstum
unterdrückt, sowohl bei Prostata-, Ovar- als auch bei Mamma-Ca
Patienten.
metavectum
Interaction
The classical view of biological organization is to consider the flow of information from
the genome to the transcriptome, to the proteome and then the metabolome. The
metabolome can be used to monitor changes in the genome or to measure the effects
of downregulation or upregulation of a specific gene transcript.
Invluence of Haelan951 to
VEGF Expression
Haelan951 (100 Days)
without Haelan951
37,5
Vascular Endothelial
Growth Factor
1,46
Prostata-Ca. patients n=5
0,97
0,97
Mamma-Ca. patients n=5
relative Genexpression
No alteration