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Joint session molecular pathology and trainees
Next generation pathology
A challenging diagnosis in paediatric pathology
11 September 2019 - SS13 Slide seminar - Next generation pathology
Gullo IreneCentro Hospitalar Universitário São João, Porto, Portugal
Faculty of Medicine of the University of Porto, Portugal
Angela Rita SementaIstituto Giannina Gaslini, Genoa, Italy
Co-authors:
Katia Mazzocco, Francesca Buffelli, Angelica ZinIstituto Giannina Gaslini, Genoa, Italy
Institute of Paediatric Research (IRP), Fondazione Città della Speranza, Padova, Italy
I have no conflict of interest to declare
Case report
• Four year-old boy
• Past medical history: irrelevant
• Family history: irrelevant
• Presenting problem: localised swelling of the right hand
• Magnetic resonance imaging:
Case report
• Four year-old boy
• Past medical history: irrelevant
• Family history: irrelevant
• Presenting problem: localised swelling of the right hand
• Magnetic resonance imaging:
Axial STIR (Short Tau Inversion Recovery) MR images
Case report
• Four year-old boy
• Past medical history: irrelevant
• Family history: irrelevant
• Presenting problem: localised swelling of the right hand
• Magnetic resonance imaging:
Axial STIR (Short Tau Inversion Recovery) MR images
3.3cm 4.6cm24 days
Case report
• Four year-old boy
• Past medical history: irrelevant
• Family history: irrelevant
• Presenting problem: localised swelling of the right hand
• Magnetic resonance imaging
• An incisional biopsy was performed
Microscopy
Microscopy
Microscopy
Microscopy
Differential diagnoses
Entity Epidemiological/clinical features
Infantile fibrosarcomaFirst year of life; 30-80% are congenital
Distal extremities, trunk, head and neck
Synovial sarcomaAdolescents/young adults; also older patients/children
~25% arise in the distal extremities
RhabdomyosarcomaChildren and adults. Location varies depending on subtyping
(embryonal, alveolar, spindle cell, pleomorphic, …)
Extraskeletal Ewing sarcomaAdolescents/young adults
Deep soft tissues of the extremities, paraspinal area
Rare entities related to specific molecular alterations:
‣Primitive myxoid mesenchymal tumour of infancyInfancy or early childhood
Trunk, extremities and head and neck
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
Adults (20-40 yy). Deep soft tissues of trunk and extremities
Children and adults. Bone/soft tissues of trunk and extremities
‣ Infantile NTRK-associated mesenchymal tumourEarly childhood
Distal extremities and trunk
Present case:
4 year-old boy, tumour in the right hand
Myxoid spindle- round- cell ST neoplasm
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Immunohistochemical study
Bcl2
Present case
Immunohistochemical study
INI1
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Present case
Immunohistochemical study
CD99
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Present case
Immunohistochemical study
SMA
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Present case
Entity Immunophenotype
Infantile fibrosarcomaVimentin
SMA±
Synovial sarcomaCK/EMA± (patchy/focal)
CD99±
Rhabdomyosarcoma
Desmin+
Myogenin+
MyoD1+
Extraskeletal Ewing sarcomaCD99 (diffuse membranous)
CK (rare)
Rare entities related to specific molecular alterations:
‣Primitive myxoid mesenchymal tumour of infancy Vimentin+
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CD99±
CD99±
‣ Infantile NTRK-associated mesenchymal tumour
SMA±
CD34±
S100±
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
Differential diagnoses: immunophenotype
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Present case
Entity Immunophenotype
Infantile fibrosarcomaVimentin
SMA±
Synovial sarcomaCK/EMA± (patchy/focal)
CD99±
Rhabdomyosarcoma
Desmin+
Myogenin+
MyoD1+
Extraskeletal Ewing sarcomaCD99 (diffuse membranous)
CK (rare)
Rare entities related to specific molecular alterations:
‣Primitive myxoid mesenchymal tumour of infancy Vimentin+
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CD99±
CD99±
‣ Infantile NTRK-associated mesenchymal tumour
SMA±
CD34±
S100±
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
Differential diagnoses: immunophenotype
Positive Negative
‣ Bcl2
‣ CD99
‣ INI1
Keratins
EMA
Myogenin (Myf4)
MyoD1
Desmin
SMA
CD34
S100
Present case
Differential diagnoses: molecular alterations
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
EntityMolecular alterations
Surrogate IHC biomarkers
Infantile fibrosarcoma ETV6-NTRK3 /
Synovial sarcomaSS18-SSX1
SS18-SSX2TLE1+
Extraskeletal Ewing sarcoma
EWSR1-FLI1 (85%)
EWSR1-ERG (10%)
EWSR1 fusion with other genes
FUS-ERG/FEV
/
Rare entities related to specific molecular alterations:
‣ Primitive myxoid mesenchymal tumour of infancy BCOR internal tandem duplications BCOR+
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CIC-DUX4; CIC-FOXO4
BCOR-CCNB3
DUX4+
BCOR+, CCNB3+
‣ Infantile NTRK-associated mesenchymal tumour TMP3-NTRK1; LMNA-NTRK1 NTRK1+
Differential diagnoses: molecular alterations
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
EntityMolecular alterations
Surrogate IHC biomarkers
Infantile fibrosarcoma ETV6-NTRK3 /
Synovial sarcomaSS18-SSX1
SS18-SSX2TLE1+
Extraskeletal Ewing sarcoma
EWSR1-FLI1 (85%)
EWSR1-ERG (10%)
EWSR1 fusion with other genes
FUS-ERG/FEV
/
Rare entities related to specific molecular alterations:
‣ Primitive myxoid mesenchymal tumour of infancy BCOR internal tandem duplications BCOR+
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CIC-DUX4; CIC-FOXO4
BCOR-CCNB3
DUX4+
BCOR+, CCNB3+
‣ Infantile NTRK-associated mesenchymal tumour TMP3-NTRK1; LMNA-NTRK1 NTRK1+
Molecular study – FLUORESCENT IN SITU HYBRIDISATION
EWSR1 break-apart SS18 break-apart
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
EntityGene fusions / other molecular alterations
Present case(RT-PCR)
Infantile fibrosarcoma ETV6-NTRK3 Negative
Synovial sarcomaSS18-SSX1
SS18-SSX2Negative
Extraskeletal Ewing sarcoma
EWSR1-FLI1 (85%)
EWSR1-ERG (10%)
EWSR1 fusion with other genes
FUS-ERG/FEV
Negative
Rare entities related to specific molecular alterations:
‣ Primitive myxoid mesenchymal tumour of infancy BCOR internal tandem duplications Negative
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CIC-DUX4; CIC-FOXO4
BCOR-CCNB3
Negative
Positive
‣ Infantile NTRK-associated mesenchymal tumour TMP3-NTRK1; LMNA-NTRK1 Not available
Molecular study - REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION
WHO classification of Tumours of Soft Tissue and Bone. 4th Edition . Fletcher C et al Eds. 2013. IARC.
Practical Soft Tissue Pathology: A Diagnostic Approach. Second Edition. Chapter 8. Hornick J Ed. 2019. Elsevier
EntityGene fusions / other molecular alterations
Present case(RT-PCR)
Infantile fibrosarcoma ETV6-NTRK3 Negative
Synovial sarcomaSS18-SSX1
SS18-SSX2Negative
Extraskeletal Ewing sarcoma
EWSR1-FLI1 (85%)
EWSR1-ERG (10%)
EWSR1 fusion with other genes
FUS-ERG/FEV
Negative
Rare entities related to specific molecular alterations:
‣ Primitive myxoid mesenchymal tumour of infancy BCOR internal tandem duplications Negative
‣ “Ewing-like sarcomas”:
- Sarcoma with CIC-related gene fusions
- Sarcoma with BCOR-related gene fusions
CIC-DUX4; CIC-FOXO4
BCOR-CCNB3
Negative
Positive
‣ Infantile NTRK-associated mesenchymal tumour TMP3-NTRK1; LMNA-NTRK1 Not available
Molecular study - REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION
Diagnosis
BCOR-CCNB3 fusion-positive sarcoma
Diagnosis
BCOR-CCNB3 fusion-positive sarcoma
Pierron G et al. Nat Genet 2012
Intra-chromosomal para-centric inversion on chromosome X resulting in
fusion between the BCOR (encoding the BCL6 co-repressor) and CCNB3
(encoding the testis-specific cyclin B3) genes
Diagnosis
BCOR-CCNB3 fusion-positive sarcoma
Pierron G et al. Nat Genet 2012; Specht K et al. Am J Surg Pathol 2016.
The term “sarcomas with BCOR-related gene fusions” has been proposed:
‣ Other BCOR gene fusion partners (MAML3, ZC3H7B and KMT2D)
‣ Sarcomas displaying BCOR rearrangements with unknown partners
Intra-chromosomal para-centric inversion on chromosome X resulting in
fusion between the BCOR (encoding the BCL6 co-repressor) and CCNB3
(encoding the testis-specific cyclin B3) genes
Sarcomas with BCOR-related gene fusions
Transcriptomic profiling
Kao YC et al. Am J Surg Pathol 2017; Pierron G et al. Nat Genet 2012
Sarcomas with BCOR-related gene fusions
At a glance
- About one-hundred cases have been reported in the literature
- Affect mainly children and teenagers (2-19 years). Few cases (n=6) have been
described also in adults patients (31-44 years)
- Show striking male gender predominance (male-to-female ratio of 4.5:1)
- Evenly distributed between bone (60%) and soft tissue (40%) of trunk and
extremities. Visceral involvement (lung, kidney) is rare
- Less aggressive clinical course than Ewing sarcomas according to some reports
Puls F et al. BCOR-CCNB3 (Ewing-like) sarcoma: a clinicopathologic analysis of 10 cases, in comparison with conventional Ewing sarcoma. Am J Sure Pathol 2014;
Machado I et al. Ewing sarcoma and the new emerging Ewing-like sarcomas: (CIC and BCOR-rearranged sarcomas). A systematic review. Histol Histopathol 2016
Kao YC et al. Am J Surg Pathol 2017; Matsuyama A et al. Am J Surg Pathol 2017; Li WS et al. Histopathology 2016
TAKE-HOME MESSAGE
Sarcomas with BCOR-related gene fusions may
display morphologic heterogeneity and
immunophenotypic overlap with other entities
Kao YC et al. Am J Surg Pathol 2017; Matsuyama A et al. Am J Surg Pathol 2017; Li WS et al. Histopathology 2016
a. b. c. d. e.
TAKE-HOME MESSAGE
a. Synovial sarcoma ➔ Both are immunoreactive for TLE1
b. Primitive myxoid mesenchymal tumour of infancy ➔ Both are immunoreactive for BCOR
c. Ewing sarcoma ➔ CD99 staining pattern interpretation
d. Solitary fibrous tumour ➔ Diffuse nuclear positivity for STAT6
e. Vascular tumours ➔ Staining for vascular markers
Sarcomas with BCOR-related gene fusions may
display morphologic heterogeneity and
immunophenotypic overlap with other entities
Kao YC et al. Am J Surg Pathol 2017; Matsuyama A et al. Am J Surg Pathol 2017; Li WS et al. Histopathology 2016
a. b. c. d. e.
TAKE-HOME MESSAGE
a. Synovial sarcoma ➔ Both are immunoreactive for TLE1
b. Primitive myxoid mesenchymal tumour of infancy ➔ Both are immunoreactive for BCOR
c. Ewing sarcoma ➔ CD99 staining pattern interpretation
d. Solitary fibrous tumour ➔ Diffuse nuclear positivity for STAT6
e. Vascular tumours ➔ Staining for vascular markers
The identification of specific
molecular alterations (FISH, RT-
PCR, RNA seq, NGS) is essential to guarantee diagnostic accuracy
Sarcomas with BCOR-related gene fusions may
display morphologic heterogeneity and
immunophenotypic overlap with other entities
Gullo Irene, MD, PhDCentro Hospitalar Universitário São João, Porto, Portugal
Faculty of Medicine of the University of Porto, Portugal
A challenging diagnosis in paediatric pathology
Sarcoma with BCOR-CCNB3 gene rearrangement
11 September 2019 - SS13 Slide seminar - Next generation pathology
Thank you for your attention
Kao YC et al. Am J Surg Pathol 2016; Kao YC et al. Am J Surg Pathol 2017; Practical Soft Tissue Pathology: A Diagnostic Approach, Second Edition. Chapter 8. Hornick J
Ed. 2019. Elsevier; Matsuyama A et al. Am J Surg Pathol 2017
BCOR immunoreactivity CCNB3 immunoreactivity
Sensitivity 100% ~90%
Other tumours showing
immunoreactivity
PMMTI
CCS kidney
SS (>50%)
Rare: ES, ERMS
Rare: SFT, ES, ERMS, fibrous
sarcoma
Surrogate biomarkers for BCOR-CCNB3 translocation
% of positive cases
BCOR 100%
CCNB3 >90%
Bcl2 >90%
CD99 ~50% - patchy/focal and not membranous (cytoplasmic/dotted)
CD117 ~50%
CD56 >90%
Cyclin D1 >90%
TLE1 80-100%
CD34 0%
SMA 0%
Desmin 0%
S100 0%
CK 0%
EMA 0%
Immunophenotypic profile of BCOR-CCNB3 positive sarcoma