An update on

immunohistochemical markers

in mesenchymal neoplasms

By Konstantinos Linos MD, FCAP, FASDP

Assistant Professor of Pathology

Geisel School of Medicine at Dartmouth

Dartmouth-Hitchcock Medical Center

Lebanon, NH, USA

• Book Royalties

Financial disclosures

A general truth!

• Some of these markers may prove to be more useful in clinical practice than others

• With time it is generally appreciated that significant overlap in staining patterns can be seen in different tumor types, some of which share similar biology or can be explained by known biologic mechanisms

• ERG• PAX7• DUX4/ETV4• BCOR• Claudin 4

Outline

ERG

• Member of the ETS family of regulatory transcription factors with diverse biological roles • Regulates endothelial cell differentiation,

angiogenesis, and expression of several endothelial-specific antigens

• Also required for embryonic stem cells to differentiate into endothelial cells

• Regulatory gene of cartilage skeletogenesis• May have crucial role in permanent cartilage

development

ERG in benign vascular tumors

ERG in

Hemangioendotheliomas

ERG in angiosarcoma in

comparison with CD31

ERG in non epithelial Mesenchymal,

Neuroectodermal and Hematopoietic

tumors

ERG in Epithelial Neoplams

ERG in nonvascular tumors

A hemorrhagic poorly

differentiated carcinoma

simulating angiosarcoma

ERG

• From these studies ERG is positive in >95% of angiosarcomas, with a greater sensitivity

compared to CD31 and CD34

• ERG usually shows a diffuse pattern of nuclear staining, which facilitates its interpretation in this context.

Ewing Sarcoma

with EWSR1-

ERG

FLI1

ERG

Ewing sarcoma with EWSR1-FLI1

EWSR1, FLI1, ERG and their

fusion proteins in Ewing Sarcoma

Leukemia cutis cases

Reactive Leukocytic Infiltrates

Soft Tissue Chondroma

Convetional Chondrosarcoma

ERG

FLI1 ERG

Conclusion

• ERG is therefore a useful marker for confirming endothelial differentiation in both benign and malignant neoplasms

• Potentially useful marker to distinguish Leukemia Cutis vs reactive myeloid infiltrates

• Expression can also be seen in a subset of epithelioid sarcomas and a small percentage of Ewing sarcomas, as well as approximately 45% to 50% of prostatic carcinomas.

• ERG can be seen in selected bone and soft tissue tumor with cartilaginous differentiation

ERG

• ERG• PAX7• DUX4-ETV4• BCOR• Claudin 4

Outline

• Immunohistochemical detection in tumor cell nuclei of myogenin and MYOD1 currently benchmark for rhabdomyosarcoma diagnosis

• Sensitivity and specificity even in combination is imperfect

• Negative or focal myogenin in a considerable number of embryonal rhabdomyosarcomas

• MYOD1: high background and cytoplasmic staining

PAX7

• PAX7 is a paired box transcription factor required for mammalian skeletal muscle stem cells (aka satellite cells)

• It plays a critical role in mammalian myogenesis

• Controls early lineage specification, whereas MYOG and MYOD1 regulate subsequent lineage commitment

PAX7

Embryonal Rhabdomyosarcoma

Embryonal Rhabdomyosarcoma

Alveolar Rhabdomyosarcoma

PAX7 and Rhabdomyosarcoma

• Potential diagnostic value of PAX7 IHC in the evaluation of rhabdomyosarcoma, especially embryonal rhabdomyosarcoma

• Less sensitive in ARMS • Could be used secondarily in desmin+,

MYOG/MYOD1 - cases

Ewing Sarcoma

CIC-DUX4

• PAX7 a sensitive marker for Ewing Sarcoma

• Positive in NKX2-2 negative cases

• Also positive in both common and variant forms of Ewing Sarcoma

• PAX7 expression differentiates Ewing Sarcoma from CIC-DUX4 sarcoma

What accounts for the robust PAX7

expression in Ewing Sarcoma?

EWSR1-FLI fusion protein binds at position 5 to the PAX7 promoter

Undifferentiated small round

cell or spindle-cell tumor

• PAX7 positivity: Rhabdo vs Ewing Sarcoma• Positivity for desmin, MYOG, MYOD1 would

strongly favor rhabdomyosarcoma• Rare cases of ES can express myogenic features

• Diffuse membranous CD99 would favor ES• Can be positive in rhabdomyosarcomas

• NKX2.2 would support ES• WT1+, PAX7- would support CIC-DUX4

• To date IHC focused on the aberrant expression of one of the genes involved in the pathogenic translocation• WT1 in DSMRCT (EWSR1-WT1)• STAT6 in SFT (NAB2-STAT6)• FLI1 in Ewing Sarcoma (ESWR1-FLI1)• BCOR in undifferentiated sarcomas

• PAX7 in ES exploits a characteristic transcriptional read-out of the disease-

causing translocation

• ERG• PAX7• DUX4 & ETV4• BCOR• Claudin 4

Outline

• Emerging undifferentiated sarcomas resembling but distinct from Ewing Sarcoma• t(4;19) translocation involving CIC-DUX4• t(10;19) translocation CIC-DUX4L

• Distinct transcriptional signature with poor clinical outcomes

DUX4 IHC

CD99 WT1 FISH CIC

Ewing Sarcoma Malignant Rhabdoid

Tumor

Synovial Sarcoma

• ETV4 (ETS Variant 4)• Member of the PEA3 subgroup in the ETS

transcription factor family • Upregulation of WT1 and ETV1/4/5 in CIC-

rearranged sarcomas

ETV4 and round cell sarcomas

• Sensitive but not entirely specific for CIC-rearranged sarcomas

• Diffuse, moderate-to-strong expression in ~90% sensitive and 95% specific

• ERG• PAX7• DUX4 & ETV4• BCOR• Claudin 4

Outline

BCOR (BCL-6 interacting corepressor)

• A subset of undifferentiated round cell sarcomas:• BCOR-CCNB3• BCOR-MAML3• ZC3H7B-BCOR• YWHAE-NUTM2B• BCOR internal tandem duplication (ITD)

• Clear cell sarcoma of the kidney• Primitive myxoid mesenchymal tumor of

infancy

BCOR and Synovial Sarcoma

BCOR

• BCOR IHC is highly sensitive in identifying sarcomas with BCOR abnormality• Triage for further molecular tests• Avoid extensive IHC and molecular workups in

small specimens• Synovial sarcoma should be included in the

DDx of round cell sarcomas with BCOR+

• ERG• PAX7• DUX4 & ETV4• BCOR• Claudin 4

Outline

Claudin 4 and SWI/SNF

complex-deficient tumors

• Claudins are integral components of tight junctions which for barriers in epithelial, endothelial and perineurial cells

• Claudin 4 is expressed in most epithelial cells and carcinomas

• Has been validated as a useful marker in the distinction of mesothelioma (lack of expression) from metastatic adenocarcinoma (consistently +)

• It has been observed:• Epithelial component of biphasic synovial

sarcoma• Subset of Desmoplastic Small Round Cell Tumor

Claudin 4

• Fundamental role in regulation of gene expression

• Tumor suppressor properties• SMARCB1 (INI-1)• SMARCA2 (BRM)• SMARCA4 (BRG1)• ARID1A

SWI/SNF complex

• SMARCB1 deficiency (INI1 loss)• Malignant Rhabdoid Tumor• Epithelioid Sarcoma• Subset of myoepithelial carcinomas• Epithelioid MPNST• Epithelioid Schwannoma

• SMARCA4 deficiency (BRG1 loss)• Nearly all cases of ovarian small cell carcinomas

of hypercalcemic type• ARID1A deficiency

• ~half of ovarian clear cell carcinomas

SWI/SNF complex

Epithelioid

Sarcoma

Epithelioid

Angiosarcoma

Epithelioid

MPNST

Biphasic

Synovial

Sarcoma

Ovarian

Clear Cell

Carcinoma

Pancreatic

Rhabdoid

Carcinoma

Cecal

Rhabdoid

Carcinoma

INI1-

deficient

Sinonasal

Carcinoma

Claudin 4

• It may serve as a useful diagnostic adjunct in the distinction of SWI/SNF complex deficient carcinomas from sarcomas with epithelioid morphology

• Strongly associated with true epithelial differentiation

• However absence of claudin 4 staining does not entirely exclude carcinoma

Keratin CD34

SMARCA4

NEVER FORGET!

• The use of all of these markers requires careful clinical correlation and knowledge of the spectrum of staining in other tumor types, as no one marker is 100% sensitive or specific

for a given diagnosis

• Email: Konstantinos.Linos@hitchcock.org

• @ @ KonstantinosLin

mailto:Konstantinos.Linos@hitchcock.org