TE pancréatique fonctionnelle : insulinome

NEUROENDOCRINE TUMORS
Diagnosis, etiopathogenesis and molecular biology
Philippe RUSZNIEWSKI
I GETNE EDUCATIONAL MEETING
Barcelona 7th April 2010
Pôle des Maladies de l’Appareil Digestif
Gastroentérologie-Pancréatologie,
Hôpital Beaujon, Université Paris VII, Clichy
Beaujon Hospital group
for the study of NEuroendocrine Tumours
(NETs) of the digestive tract
French Referent Center for the study of NETs
Gastroenterology-Pancreatology:
Pr P. RUSZNIEWSKI (ENET Vice President), Pr P. HAMMEL, Dr O. HENTIC
Oncology:
Pr E. RAYMOND, Pr S. FAIVRE
Surgery:
Pr A SAUVANET, Dr S DOKMAK, Pr BELGHITI
Pathology:
Dr A. COUVELARD
Imaging:
Pr V. VILGRAIN, Dr M. ZAPPA
Genetic:
Dr D. VIDAUD
Where are NETs localized ?
GI tract: 75%
Stomach (10%), Pancreas (10%)
Small bowel (30%)
Rectum (20%),
Appendix (20%)
Colon (<5%)
Plöckinger, Neuroendocrinology 2004
Modlin, Cancer 2003
Broncho-pulmonary: 25%
NET are rare tumours… but their incidence is increasing !
(Survey, Epidemiology and End Results (SEER), US population 1974–2005) 2.5–5.0/100,000 Inhabitants)
•
Data from the US SEER database indicate an approximate 5-fold increase
in the incidence of NETs between 1975 and 2004
1,4
Per 100,000 Population
1,2
1,0
0,8
0,6
NET Site
Lung
Appendix
Stomach
Colon
Small Intestine
Rectum
Caecum
Pancreas
0,4
0,2
19
73
19
74
19
75
19
76
19
77
19
78
19
79
19
80
19
81
19
82
19
83
19
84
19
85
19
86
19
87
19
88
19
89
19
90
19
91
19
92
19
93
19
94
19
95
19
96
19
97
19
98
19
99
20
00
20
01
20
02
20
03
20
04
0,0
This increase is most likely due to improved detection such as
the availability of advanced radiological and endoscopic imaging.
Yao JC, et al. J Clin Oncol 2008;26:3063–3072
Prevalence of different tumours in the US (2004)
120 000
103 312
Diagnosis of NET :
Endoscopy performed for
unrelated symptoms
100 000
Prevalcence
80 000
65 836
60 000
40 000
32 353
28 664
21 427
20 000
0
NET
Gastric
Pancreatic
Neoplasia
Oesophageal
HCC
Epidemiology of pancreatic NET (pNETs)
• Autopsy : 1.6 to 10%per year
• Clinical incidence : 2- 4 per million/year
• 2%-10% of pancreatic tumours
• Prominently non-functional (2/3)
• Equal distribution male/female
• Peak incidence in individuals aged 50 years
Kimura W, Dig Dis Sci 1991
Barakat MT, Endocr Relat Cancer 2004
Mignon M, Digestion 2000
Criteria for assessing prognosis of NET
•
WHO classification
(ENET consensus, Neuroendocrinology 2006, 84:151-216)
Tumour size Angio- Proliferation
Metastases
Differentiation
/ Invasion
(cm)
invasion
Ki67 %
Behavior
WHO
Benign
(low risk)
Group 1
-
Well-D
≤2
-
<2
Benign
Group 2
(Intermediate risk)
-
Well-D
>2
+/-
~2
Malignant
Low-grade
Group 3
+
Well-D
carcinomas
>3
+
>2
Malignant
High-grade
Group 4
+
Poorly-D
carcinomas
any
+
> 20
Differentiation of NETs
Well-differentiated
Poorly-differentiated
Epidemiology of pancreatic NET (pNETs)
Rate
80%
20%
Secretion
Yes
No
Usually low
High
Good
Bad
Proliferation
Prognosis
Differentiation of pNETs & survival
Metastic Tumour (Stage 4)
5-year Survival Rate (%)
4
NET: poorly differentiated1
>35
NET: well/moderately differentiated1
1.2
56*
Rate of survival (%)
1.0
Tumour with good differentiation (18 of 64 died)
33*
17*
0.8
11*
8*
0.6
4*
0.4
3*
Tumour poorly differentiated (7 of 9 died)
0.2
1*
0*
24
36
0
0
12
48
60
72
Follow up (months)
Madeira I, Gut 1999
84
96
108
120
In what conditions are NETs diagnosed ?
Symptoms due to
Hormonal hypersecretion
By accident ++
Symptoms caused
by mass
(imaging or endoscopy)
Diagnosis
Systematic screening
(MEN1)
Incidence of PETs is increasing (1)
24 Italian Centers June 2004-March 2007
297 new cases with sporadic PET
(75% from surgical centers)
58.6 years
51% Female (F : 63%, NF : 48%)
75.4% of PETs were NF, half discovered incidentally
F-PETs : insulinomas 73%, gastrinomas 20%
Am J Gastroenterol 2009; 104:3034-41
Comments
• Family history of cancer (pancreatic adk)
• Chronic pancreatitis
• High alcohol intake
• Recent-onset diabetes
• (Smoking)*
→ Overlap between endocrine and exocrine
pancreatic carcinogenesis ?
* Significant at univariate analysis only
Diagnosis of NETs: hormonal secretion
• Chromogranin A: soluble secretory
glycoprotein
– general marker (in 60%-80% of NETs)
– useful in non-functioning tumours
– false positive:
• renal failure
• hyperplasia of ECL cells (PPI, atrophic gastritis)
ECL : enterochromaffin-like
Functioning pNETs : secretions
•
•
Hormone
Consequence
Gastrin
Acid : ulcers, diarrhoea
Insulin
Glucagon
VIP
Somatostatin
Hypoglycaemia
Hyperglycemia (necrolytic migratory erythema)
Cholera-like diarrhoea
Biliary lithiasis
Zollinger-Ellison syndrome
Other secretions by functioning pNETs: calcitonin, PTH-RP, GH-RH…
Intestinal NETs – but not pNETs - produce serotonin & tachykinins
causing diarrhoea and flushes
VIP : vasointestinal peptide, PTH-RP : parathormon-related peptide, GH : growth hormone
Marqueurs généraux : chromogranine A
•
Glycoproteine de structure des granules de sécrétion
Seul marqueur à doser systématiquement
•
Le + sensible quel que soit le primitif
•
Sensibilité
- si tumeur volumineuse
- si tumeur sécrétante
- selon l’origine embryologique
Intestin antérieur (sauf insulinome) > moyen > postérieur (rectum)
- selon la différenciation
•
Marqueur de progression tumorale ? non validé : VPP 24%
Vezzosi et al, JFHOD 2009 CO 105
Marqueurs spécifiques
□ Selon le syndrome hormonal …..et non….
□ TE pancréatique fonctionnelle
- insuline
- gastrine
- VIP
- glucagon
- somatostatine
□ TE intestin grêle fonctionnelle
- 5HIAA urinaire
TE pancréatiques fonctionnelles
Diagnostic
Signes cliniques
Biologie
Epreuve
dynamique
Insulinome
Triade de
Whipple
Hypoglycémie
Hyperinsulinémie
Epreuve de
jeûne
Zollinger-Ellison
Hypergastrinémie
Débit acide basal ↑
Test à la
sécrétine
Gastrinome
Ruszniewski et al. Spectre clinique des tumeurs neuro-endocrines. Revue du praticien 2002. 52:19
TE pancréatiques fonctionnelles
Diagnostic
Signes cliniques
Biologie
Epreuve
dynamique
Verner Morrisson
HyperVIPémie
(VIP= Vasoactive
intestinal peptide)
Epreuve de
jeûne
négative =
diarrhée
sécrétoire
VIPome
Glucagonome
Hyperglucagonémie
Erythème nécrolytique
migrateur, diabète, cachexie
Somatostatinome
Lithiase vésiculaire,
stéatorrhée
Hypersomatostatinémie
Ruszniewski et al. Spectre clinique des tumeurs neuro-endocrines. Revue du praticien 2002;52:19
TE pancréatique fonctionnelle : insulinome
- Sécrétion inappropriée d’insuline : Epreuve de jeûne
- Après interrogatoire +++ :
-
hypoglycémies (< 0,4-0,5 g/l) avec signes adrénergiques et/ou
neuroglycopéniques
à jeun ou à l’effort
avec efficacité du re-sucrage et
éliminer prises médicamenteuses
TE pancréatique fonctionnelle : insulinome
•
Epreuve de jeûne
-
en hospitalisation pendant 72 h avec perfusion de G10% branchée en Y
surveillance clinique +++
dextro / 6 h et en cas de « symptômes d’hypoglycémie »
interruption si < 0,40 g/l ( 2,2 mmol/l) et / ou troubles neuropsychiques
aigus
confirmation par glycémie veineuse et dosage insulinémie, pro-insuline et
peptide C
-
TE pancréatique fonctionnelle : insulinome
- Interruption car positive
30 % à 12 h
70 % à 24 h
90 % à 48 h
100 % à 72 h
Sécrétion d’insuline (et de pro-insuline) inadaptée à l’hypoglycémie et
concentration non freinée de peptide C
≠ hypoglycémie factice
TE pancréatique fonctionnelle : gastrinome
•
Gastrinémie
•
•
Faux positifs +++
Secondaires à une hypo ou achlorydrie ( = peu ou pas d’acide)
-
IPP (arrêt 7 à 10 jours)
gastrite fundique atrophique (type Biermer)
insuffisance rénale
sténose duodénale
SI doute diagnostique : test à la sécrétine
TE pancréas fonctionnelle : gastrinome
Se %*
Spe % **
DAB
> 31 mmoles/h
50
99
DAB sous
sécrétine
> 18 mmoles/ h
78
100
Gastrinemie
basale
> 325 pg/ml
53
100
Gastrinémie
sous sécrétine
> 270 pg/ml
80
100
* % de pts avec gastrinome au dessus de la valeur seuil
** % de pts avec UD au dessous de la valeur
TE intestin grêle fonctionnelle : carcinoïde
Pas de dosage de la sérotonine plasmatique ou urinaire ( trop peu spécifique)
•
•
•
Dosage de son métabolite urinaire: 5 HIAA (acide 5- hydroxyindole
acétique) 3 jours de suite
Recueil des urines dans flacon opaque avec 20 ml d’HCL
Après 3 jours de régime pauvre en tryptophane (chocolat, banane,
avocat, fruits secs…)
Se 80%, Spe 95%
Gene expression profiling
in GEP-NET
Pancreatic-ET
Maitra
2003 Clin Cancer Res
PET vs microdissected islets
Hansel
2004 Clin Cancer Res
PET: 5 metastases- vs 7 metastases+
Durkin
2004 J Am Coll Surg
PET vs normal / ductal K (n=27)
Bloomston 2004 Annals Surg Oncol PET vs normal / pancreatitis / K (n=9)
Dilley
2005 Molecular Cancer
PET-MEN1 (n=8) vs ilets (n=4)
Couvelard 2006 Gastroenterology
PET : 12 benign vs 12 WD carcinomas
Capurso
2006 End Relat Cancer
PET : 8 prim vs 5 mét vs 4 islets
Duerr
2008 End Relat Cancer
PET ≠ stages (n=24) and GI-NET (n=6)
PET ≠stages
Normal counterpart
Ileal-ET
Duerr
2008 End Relat Cancer
GI-NET (n=6) and PET ≠ stages (n=24)
Arvidsson 2008 End Relat Cancer
GI-NET (n=5) / cell cultures (BON; GOT1)
Leja
GI-NET: 3 metas- vs 3 metas+ / normal
mucosa
2009 Modern Pathology
Normal counterpart
Bloomston, Annals Surg Oncol, 2004
They compared PET with normal pancreas and found genes involved in
different pathway, including ANG2 which could play a role in the
angiogenesis and progression of PET
PET vs normal, pancreatitis, ductal carcinomas
Pooled sample expression profiling (9 samples pooled in each group)
Affymetrix microarrays
IHC
Angiopoietin 2 +++ in PET
Control Q-RT-PCR
Importance of angiogenesis,
remodelling
NPDC1: neural proliferation, differentiation, and control; ANG2: angiopoietin 2; ELOVL4: Elongation of very-long-chain fatty
acids 4-like protein; CALCR: calcitonin receptor
Bloomston, Annals Surg Oncol, 2004
Couvelard, Gastroenterology, 2006
Profile of benign vs malignant PET, identified genes involved in different
pathway (angiogenesis, signal transduction/tyrosine kinases, calciumdependent cell signaling, response to drug)
•
cDNA microarrays (Sanger
Center, UK) with 9932 cDNA*
•
Validation
– qRT-PCR
– Tissu-array (n=129 PET)
•
Patients and tumours
–
•
Classified according to WHO:
–
•
24 surgically resected PET
12 benign and 12 carcinomas
Data recorded:
–
–
–
–
–
VHL disease
tumour size
MIB1 index
microvascular density
metastasis
WHO
benign
benign
benign
benign
benign
uncertain
uncertain
uncertain
uncertain
uncertain
uncertain
uncertain
VHL
0
1
0
0
0
0
0
0
0
1
0
0
size (mm)
13
20
9
20
18
25
23
28
25
45
35
21
lymph node
0
0
0
0
0
0
0
0
0
0
0
0
liver metastasis
0
0
0
0
0
0
0
0
0
0
0
0
MVD>200
1
1
1
1
1
1
1
1
1
1
1
1
MIB %
1
2
1
2
1
1
1
2
4
3
6
6
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
carcinoma
0
1
0
0
1
1
0
0
0
0
0
0
70
30
50
30
22
30
100
20
160
100
43
50
1
1
1
1
1
1
0
0
1
1
1
1
0
0
0
0
0
0
1
1
1
1
1
1
1
1
1
1
1
0
0
0
0
0
0
0
2
3
1
1
4
5
6
7
8
10
20
10
*http://www.sanger.ac.uk/Projects/Microarrays
Results
M
133 transcripts differentially regulated
between benign and malignant tumours
79 up- and 54 down-regulated in
metastatic tumours
M
B
B
Different classes/pathways
– Angiogenesis and remodeling
• CD34, cadherin-5, E-selectin, semaphorin E, and
fibrillin)
– Signal transduction / tyrosine kinases
• tyrosine kinase-2, IGF1, PDGFR-β, MKK4,
discoidin domain receptor-1
– Calcium-dependent cell signaling (hormonal
release and vesicular trafficking)
• transient receptor potential cation channel-1,
calcium channel voltagedependent 2, neurocalcin,
GABA-A receptor 2
– Response to drug
• MDR1, CEA–related CAM 6
*
* IGF signalling; Hansel DE; Clin Cancer Res 2004
Confirmation by immunostaining on tissu-array including 110 PET
Benign
E-selectin: endothelial adhesion molecule,
trafficking and angiogenesis, immune-mediated
response to tumor
MKK4: mitogen-activated protein kinase
kinase 4
E-selectin
MDR1: multidrug resistance 1; ATP-dependent
drug efflux pump; mediates the development of
resistance to anticancer drugs.
MKK4
MDR1
Malignant
Ileal ET
– Clustering:
• I-ETs cluster separately from PETs
– Comparison malignant P-ET (n=19)
vs malignant I-ET (n=6)
• 385 genes upregulated in I-ET
• involved in ion, channel and
neurotransmitter transport were
significantly overrepresented in I-ET
• 3 most highly up-regulated genes in I-ET
– VMAT1: vesicular monoamine member1
– ECM1: extracellular matrix protein1
– Galectin 4
– Explored PDGFRβ and RET
Duerr, Endocr Related Cancer, 2008
Conclusion
•
Microarray analysis performed in different subtypes ot NET can give
some informations:
– Molecular profiling demonstrate that P- and I- ET display different genetic
changes
– Interesting set of validated molecules (APP…)
•
Dat are numerous and the investigator must choose among many
potentially interesting genes for further studies++
– to determine the role of newly discovered molecules in tumorigenesis
– To better understand their function
•
No significant overlap between genes in the different studies!!! (some
genes: IGF1/APLP1++/SERPINA10/GADD45b), and this may due to
different study design??
–
–
–
–
Platforms?
Sample subtypes? Including MEN1/VHL; functional ±; malignant ±
Sample size? From 3 to >15 in each group
Comparison of groups? benign vs malignant ; tumour vs normal
counterparts ; cell culture
• Involved in different pathways
– Multidrug resistance pump (MDR1): cellmembrane transporter; efflux of drugs
– Thymidylate synthase (TS): interferes with CT
metabolites (5-FU)
– Apoptosis (Akt, PTEN, p53, bcl2, Ki-67):
modify drug induced apoptosis
– Mismatch repair (hMLH1): influence response
through interaction with cell cycle regulation
– Hypoxia & vascular regulating mechanisms of
drug resistance (drug delivery): HIF1, CA9,
CD34, VEGF
Patients
• 60 patients with complete f/u data
• Progressive and advanced/metastatic
GEP-NETs
– Pancreas 50%
– Ileum + colon 25%
– Other locations ….
• Histopathological data (differentiation,
grade according to TNM classification)
SUMMARY
Association between markers and CT
response
5FU
MDR1
DOX
Mib-1
p53
VP16/CDDP
STZ