Transcript Slide 1

Oncogenic Mutation Screening
in Solitary Fibrous Tumors
Elizabeth G. Demicco, Khalida Wani, Kenneth
Aldape, Alexander J. Lazar, and Wei-Lien Wang
Solitary Fibrous Tumor (SFT)
Background
Phenotypic spectrum comprising classic solitary fibrous tumor
(hypocellular variant) and hemangiopericytoma (hypercellular
variant)
Rarely metastasizing– Behavior can be difficult to predict
Solitary Fibrous Tumor (SFT)
Background
Limited data on cytogenetic abnormalities.
Relatively simple genomic alterations
Few recurrent features
Limited data on genetic abnormalities
PDGFRB mutations, 2/88 pleural SFT
No mutations in DDR1 (n=8), ERBB2 (n=10), FGFR1
(n=15), PDGFRB (n=39)
TP53 one case, dedifferentiated SFT of nasal cavity
Purpose
Examine a large series of solitary fibrous
tumors for common oncogenic mutations.
Screening Approach to Mutational Analysis
(Sequenom MassArray)
Detection method for single nucleotide
polymorphisms (SNP)
Step 1. PCR-based
Allele (WT vs. mutant) specific probes
Single nucleotide extension across site of SNP
Different probe sizes allow for allele differentiation
and multiplexed reaction
Step 2. MALDI-TOF mass spectrometry
Analyze primer extension product
Time of flight mass spectrometry differentiates
based on probe size
Gene
# Sites
Alleles
Gene
# Sites
Alleles
AKT1
1
G49A
CTNNB1
12
A95CGT; G94CAT, G101ATC; A107CG; T104AGC; T97GCA;
T109GCA; C110GTA; T133GCA; C134GTA; A121GCT; C122TAG
AKT2
1
G49A
KIT
12
AKT3
1
G49A
A2447GTC; G2446CAT; A1924G; T1727C; A1696G; T2466GCA;
A2464TCG; C1900T; T1676CAG ; T1679AGC ; T2474C ; T1657A
BCOR
1
A4220GCT
BRAF
12
CC2D1A
1
C3036G
A1781GT; G1756A; G1391ATC; G1397ATC; G1396CA; G1406ATC;
G1405CA; A1801G; T1790GA; G1800ATC; G1800ATC; T1799ACG_F;
T1799ACG_R; G1798TA
EPHA3
1
G2283TC
GNAS
2
G602A; C601AT
FOXL2
1
C402G
PIK3CA
28
GRM3
1
G2608A
JAK2
1
G1849T
C3137T; T1258C; G328A; G1252A; G1357A; G1624AC; A1625TG;
A1634CGT; G1635CT; G1633AC; C2727G; G353A; A3140GT_F;
A3140GT_R; C3139T; A2102C; G333C; G3129ATC; A3127G; T1035A;
C1616G; C178A; C1636GA; A1637TCG; G263A; C1214T; A3073TG;
A3062G; T3061CA
MGA
1
C5421A
IDH1
3
G209A; C394TGA; G395AT
PPP2R1A
1
T769G
IDH2
4
G419TA; A514GT; G515TA; G516T
RET
1
T2753C
MAP2K7
4
C870T; G485A; T811A; C932T
RPL22
1
A44CGT
PDGFRA
5
A2525T; G2524TA; C1977A; A1975T ; T1682A
SFRS9
1
C722A
FBXW7
6
C1393T; G1394AT; G1436AT; C1513TA; G1514ATC; C1745T
SMO
1
G970A
RAF1
6
G955T; C1837G; A344G; G1005C; T775G; T1018G
SRC
1
C1591T
FGFR3
7
G1108T; G1138A; G2089T; A1949TC ; C742T ; C746G ; A1118G
TGM2
1
T734C
KRAS
7
G436CA; G28A; G35ACT; G34ACT; G38ACT; G37ACT; C181GAT
CDK4
2
C70T; G71A
MET
2
G1225T; C9013T
7
A3335GT; C3334T; T3803C; A1124G ; C2962T ; C3029T ; T3742CG
CSMD1
FBXO4
2
T68A; C226A
ALK
8
T3521GC; C3522AG; T3520CAG; T3734G; C3735AG; T3733GA;
T3512A; G3824AT
FGFR1
2
C754A; C374T
EGFR
8
G2155TA; A2579T; T2573G; T2582AG; T2158C ; C2369T ; C2561T ;
A2438G
FGFR2
2
T1647GA; C755G
NRAS
8
GNA11
2
A626TC; C547T
G436A; G35ACT; G34ACT; G38ACT; G37ACT; C181GAT; A183TCG;
A182GCT
GNAQ
2
A627T; A626TCG
Tumor Characteristics
122 tumors (105 patients)
72 Primary
9 Local recurrence
41 Metastasis
13 Patients with multiple tumors tested
6 primary and metastasis(es)
5 multiple metastases
2 local recurrence(s) +/- metastasis
Patient Demographics
Site
Number of
Patients
Gender
(% F)
Age at first diagnosis, years
(median, range)
All
105
48.5%
53, 16-89
Meningeal
24
38.9%
45, 16-89
Pleural
22
36.4%
63, 35-81
Intraabdominal
27
51.9%
53, 27-71
Extremity
15
70.0%
57, 31-69
Trunk
10
53.3%
51, 29-78
Head and
neck
7
57.1%
56, 39-72
General Quality Control Indicators
Variable
Run 1
Run 2
Total
Number of cases (each in
duplicate)
31
96
127
Total number of
polymorphism sites tested
175 (41 genes)
174 (41 genes)
174/175
% Successful reads
96.5%
91.9%
93.0%
Duplicate concordance
success rate (out of all
possible reads)
94.4%
86.5%
88.4%
Duplicate failure rate due to
no result one or both
duplicates (out of all possible
reads)
5.3%
13.2%
11.3%
Duplicate discordance rate
(WT/mut read) (out of all
possible reads)
0.3%
0.3%
0.3%
Potential Mutations Identified by
Sequenom
Run 1
Run 2
Total
12
(0.23%)
21
(0.15%)
33
(0.17%)
ALK T3733G (p.F1245V)
-
1
1
BRAF T1799A* (p.V600E)
2
1
3
KRAS G35A (p.G12D)**
1
10
11
KRAS G34A (p.G12S)**
-
1
1
KRAS C181A (p.Q61K)
-
1
1
MET C2962T (p.R988C)
1
1
2
MET C3029T (p.T1010I)
-
3
3
NRAS G35A (p.G12D)**
7
3
10
NRAS G38A (p.G13D)**
1
-
1
Number SNPs detected
(% of all useable reads)
Site
* Mutation identified on forward probe, WT on reverse
** Problems with probes later reported – Determined to be WT by CAST-PCR
Characteristics of Tumors with
Identified SNPs
Patient
(phenotypic variant)
Mutation
Tumor status
Primary Site
Site this
metastasis
Outcome
SFT118
(hypercellular)
MET C3029T
(p.T1010I)*
Metastasis
(10 yr)
Intraabdominal
Peritoneum
DoD 18 yr
SFT118
(hypercellular)
MET C3029T
(p.T1010I)*
Metastasis
(14 yr)
Intraabdominal
Peritoneum
DoD 18 yr
SFT055
(hypocellular)
MET C3029T
(p.T1010I)*
Primary
Intraabdominal
Dead other
causes 30 months
SFT048
(hypercellular)
MET C2962T
(p.R988C)*
Primary
Intraabdominal
Metastasized at
40 mo.
DoD 8.5 yr
SFT092
(hypocellular)
MET C2962T
(p.R988C)*
Primary
Pleura
LTFU 1 month
SFT067
(hypercellular)
ALK T3733G
(p.F1245V)
Primary
Intraabdominal
NED 69 mo
SFT112
(hypercellular)
KRAS C181A
(p.Q61K)
Metastasis
(13 yr)
Intraabdominal
* Presence of SNP did not correlate with c-MET expression by IHC
Peritoneum
Dod 16 yrs
MET C3029T (p.T1010I) and
C2962T (p.R988C)
Also designated as T992I and R970C
Juxtamembrane domain
Initially reported as rare somatic oncogenic mutations
Identified in a wide range of tumors
Frequency in involved cancers ~1-10%
Later proposed to represent germline polymorphisms
Seen in ~1% of individuals without cancer (1/96)
Germline in several patients with cancer
No evidence of transformation in cell models
T1010I as cooperative germline oncogenic mutation?
Identified in ~4.5% familial CRC
Mutation proposed to indirectly activate c-MET via inhibition of inhibitor
Not initiator, may promote progression
ALK T3733G (p.F1245V)
Kinase-activating mutation
Rare mutation
Reported in neuroblastoma
KRAS C181A (p.Q61K)
Activating mutation reported rarely in colonic
and lung adenocarcinoma, misc. other
carcinomas various sites
Much more rare than the equivalent NRAS
mutation
Summary
We performed screening SNP analysis in a
large series of SFT
Confirmed that SFT have low frequency of
mutations in oncogenes commonly reported in
other malignancies
7/122 cases (5.7%)
Abdominal location predominant
Insufficient data on relevance to status, prognosis
Conclusions
Mutations in commonly identified oncogenes
do not appear to function in pathogenesis of
SFT.
Alternative mechanisms?
Mutations in uncommon genes
Imprinting
miRNA regulation
Studies are ongoing
Thank You.