High prevalence of SDHB mutationsin a series

of head and neckParaganglioma from Belgium

A. Persu, V. GrA. Persu, V. Gréégoire, P. Garin, H. Reychler, G. Mortiergoire, P. Garin, H. Reychler, G. MortierJ.F. De Plaen, M. Hamoir and M. Vikkula.J.F. De Plaen, M. Hamoir and M. Vikkula.

SDHB

SDHD

L. Hederstedt. Science. 2003;299:671-2A. Persu and M. Vikkula, Current HTN Reviews 2005;1:267-273

SDHC

SDH subunitsSDH subunits

Krebs Cycle

Respiratory chain

jugularjugularvagalvagalcarotidcarotidtrachealtracheal

aorticaortic

ZuckerZuckerkkandandll

adrenaladrenal

Pheochromocytoma and paragangliomaPheochromocytoma and paraganglioma

Head and neck paragangliomaHead and neck paraganglioma

Pheochromocytoma

Defraigne et al, Defraigne et al, Rev Med LiRev Med Lièègege 1997; 52:4851997; 52:485--497497

Paragangliomas(Adrenal pheochromocytoma)

Hypoxia-like gene activation pattern

Gimenez-Roqueplo et al.AJHG., 69:1186-1197, 2001

HIF2α HIF1α

VEGF

VEGF VEGF-R1

VEGF-R1

L Wayllace. L Wayllace.

ThThèèse de Doctorat, UCL 1992se de Doctorat, UCL 1992George DJ, Kaelin WG Jr.N Engl J Med. 2003;349:419-421

Link pheochromocytoma and Link pheochromocytoma and SDHSDH genesgenes??

Dahia PLM et al.. PloS Genet.2005; 1: e8

Complex IIKrebs cycle

Prolyl-hydroxylases-

First international conference on pheochromocytoma

Bornstein and Gimenez-Roqueplo Ann. N.Y. Acad. Sci. 1073: 94-103 (2006)

Frequency of mutations in apparently sporadic pheo-PGL

Prevalence of SDHD and SDHB mutations

14%

39%

50

40

30

20

10

0SDHD SDHB

% Mut Head and neck PGLHead and neck PGL(n=69)(n=69)

7%2%

50

40

30

20

10

0SDHD SDHB

% Mut PheochromocytomaPheochromocytoma(n=299)(n=299)

Bayley, BMC Medical Genetics 2005; 6:39

Genes associated with pheochromocytoma

Gene

VHL

RET

NF1

SDHB

SDHD

Chromosome

3p25-26

10q11.2

17q11.2

1p36.13

11q23

Mutations

2-11 %

< 5 %

Unknown

3-10 %

4-7 %

Malignancy

5 %

3 %

11 %

50 %

< 3 %

Pacak K et al. (2007) Nat Clin Pract Endocrinol Metab 3: 92–102

Gimenez-Roqueplo. Ann. N.Y. Acad. Sci. 107: 112-121 (2006)

Pheochromocytoma: which genes should we test?

Clinical work-up+

Family pedigree

Targeted genetic testing

Family history or other syndromic feature Sporadic presentation

NF1

VHL RET

VHL and SDHB

No mutation

SDHDSDHD SDHB

Bilateral tumor

Malignant tumor

Extra-adrenal tumor

VHL

SDHB

SDHDSDHB

Bornstein and Gimenez-Roqueplo Ann. N.Y. Acad. Sci. 1073: 94-103 (2006)

?

…..and in Belgium?

Aim of the studyAim of the study

•• To look for the prevalence and nature of To look for the prevalence and nature of SDHSDH, , RETRET and and

VHL VHL mutations in patients mutations in patients withwith paragangliomaparaganglioma andand

pheochromocytoma pheochromocytoma from Belgiumfrom Belgium

•• To detect possible genotypeTo detect possible genotype--phenotype correlationsphenotype correlations

MethodsMethodsWe recruited all patients with head and neck PGL seen in the We recruited all patients with head and neck PGL seen in the main Academic Centers in Belgium from May 2003 to main Academic Centers in Belgium from May 2003 to December 2006, as December 2006, as wwell as abdominal PC/PGL referred to the ell as abdominal PC/PGL referred to the Cliniques UniversitaCliniques Universitaiires Saint Lucres Saint Luc during the same period.during the same period.

Written informed consent, standardized clinical information Written informed consent, standardized clinical information and blood samples for DNA extraction were obtained for alland blood samples for DNA extraction were obtained for allpatients.patients.

Screening of the coding parts of Screening of the coding parts of SDHSDH gegennes, es, RETRET and and VHLVHLwas performed by SSCP and was performed by SSCP and heteroduplexheteroduplex analysis, followed analysis, followed by sequencing whenever a shift was observed. by sequencing whenever a shift was observed.

Main characteristics of familial and sporadic PC/PGL

Familial61336.3 ± 10.68 (73%)4 (36%)3 (27%)3 (27%)

Sporadic30-51.4 ± 16.13 (10%)2 (7%)4 (13%)1 (13%)

Index patientsPatients analyzedAge at discovery (y)BilateralMultipleRelapsingEctopic*: 15 adrenal PC

Sporadic18-48.1 ± 17.11 (0.6%)03 (17%)3 (17%)

hnPGL Abd.PC/PGL*

Proportion of patients harbouring mutations

Familial(n=6)

6 (100%)

6 (100%)

-

-

-

Sporadic(n=30)

13 (43.3%)

4 (13.3%)

8 (26.7%)

-

1 (3.3%)

Overall

SDHD

SDHB

RET

VHL

No mutations were found in SDHC

Sporadic(n=18)

3 (16.8%)

1 (5.6%)

1 (5.6%)

1 (5.6%)

-

hnPGL Abd.PC/PGL

P81LP81L

c.277del Tc.277del T

c.404del Cc.404del C

c.238c.238--259del 22259del 22

c.170c.170--11GG>T>T

c.170c.170--11GG>T>T

SDHD mutations(9 in 11 patients)

P81Lc.238-259del22*c.277delT c.404delC*

c.170-1G>T(n=2)

c.238-259del22*

Familial

Sporadic

11 2 3 4

H50R(n=2)

R70M

c.314 +1-G>C*

* Previously undescribed

c.541-2A>C* R230H R242H (n=3)

N248K*

c.166-70delCCTA

Sporadic

1 2 3 4 5 6 7 8

SDHB mutations(7 in 9 patients)

S163P

c.1-36 T>C*

* Previously undescribed

Prevalence of SDHD and SDHB mutationsin sporadic head and neck PGL from Belgium

27%

13%

50

40

30

20

10

0SDHD SDHB

% of hn PGLpatients with SDHD/Bmutations

X 2

Characteristics of sporadic hnPGL according tothe presence or absence of SDHB/D mutations

SDHD+(n=4)

57.3 ± 19.1

3 (75%)

1 (25%)

1 (25%)

-

SDHB+(n=8)

51.8 ± 11.2

-

1 (12.5%)

-

-

Age (y)

Bilateral

Multiple

Relapsing

Ectopic

SDHD-SDHB-(n=19)

49.3 ± 17.5

1 (5%)

-

4 (21%)

1 (5%)

Characteristics of SDHB+ head and neck PGLMutationsc.166-70delS163Pc.541-2 A>CR230HR242H""N248K

Age4473604444415949

F-UP412411442212

SiteCBJTJTJTJTXJTX

DescribedAdrenal PC1

Adrenal PC2

NoMalignant PGL3

Metastatic abd.PGL4

in father and sonDop. Secr. Bil CB5

No

Octreo?

Med?----?-

1Mora et al., Pediatr Blood Cancer. 2006; 47:785-89; 2Cascon et al., J Med Genet. 2004; 41:e30; 3Brouwers et al.,JCEM 2006; 91:4505-09; 4Young et al. JCEM 2002; 87:4101-05; 5Jeffery et al. Ann Clin Biochem. 2006; 43:156-60.

ConclusionConclusion (I)(I)

In patients with abdominal PC/PGL:

• The overall prevalence of mutations was rather lowcareful exclusion of syndromic forms?

under-referral of patients previously genotyped for RET, VHL?

• No SDHB mutation was detected mainly adrenal PC?

ConclusionConclusion (II)(II)

In patients with head and neck PGL:

• The prevalence of SDHD mutations was 100% in familialcases but low in apparently sporadic cases.

• Most SDHD mutations were located in exon 3, aspreviously reported; 4 were not described previously

• The 2 founder mutations (B92Y, L139P) at the origin ofmost familial PGL in the Netherlands were not detected.

ConclusionConclusion (III)(III)

By contrast with previous works, SDHB mutations

• were detected in 1/4 patients harbouring head and neck

PGL, i.e 2-fold more frequently than SDHD mutations

• were found in patients with unilateral hnPGL, without

evidence of ectopic, relapsing or malignant tumours.

DiscussionDiscussion

• SDHB in patients with hn PGL: additional evidenceLima et al., J Clin Endocrinol Metab. 2007 Sep 11

• SDHD in patients with malignant PGLHavekes et al., J Clin Endocrinol Metab. 2007; 92:1245-1248.

• Genotype-phenotype correlations are less clear-cutthan previously thought

• These data should be taken into account in future recommendations for genetic testing of patientswith PC-PGL

PerspectivesPerspectives

• Further genetic characterization and genotype-phenotype correlations in our database:112 samples (75 hn PGL; 37 abdominal PC-PGL)

• Molecular and enzymatic studies in available tumour samples

• Inclusion in international databases; contributionto the elaboration of recommandations.

SDH genes

Hypoxia

Endocrine HTN Cancer

Angiogenesis

The The UCL PCUCL PC--PGL Collaborative GroupPGL Collaborative Group

Shall we discover additionalpredisposing genes?

No No SDHDSDHD, , BB or or C C mutation: mutation: further genetic heterogeneity?further genetic heterogeneity?

Multiple Multiple RareRareLocalizationsLocalizations

IV.2

V.2

V.2

V.2V.2

succinate dehydrogenase activity measuredin a liver PGL sample was not significantly

decreased compared to controls

(P. Rustin, INSERM U676, Paris)

AA

BB

AB

Microchip technology

No SDHD, B or C deletionNo other deletion

Courtesy of M Amyere