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Role of ihc on soft tissue tumours
- 1. ROLE OF IHCON SOFT TISSUE TUMOURS Dr.T.Arivazhagan Post graduate Department of Pathology
- 2. Introduction SOFT TISSUE( Mesoderm) Nonepithelial tissue: Excludes: 1.Adipose tissue 1.Skeleton & joint 2.Skeletal & Smooth muscle 2.CNS 3.Blood & Lymph vessels 3.Hematopoietic 4.PNS 4.Lymphatic tissue
- 3. WHO classification ( Histogenesis) 1.Adipocytic tumours 2. Fibroblastic/myofibroblastic tumour 3. Fibrohistiocytic tumour 4. Smooth & Skeletal muscle tumour 5. Vascular tumour 6. Pericytic/perivascular tumour 7. Chondro-osseous tumour 8. Tumours of uncertain differentiation
- 4. Ancillary techniques 1. Histochemistry 2.Electron microscopy 3. Immunohistochemistry 4. Cytogenetics & Molecular studies
- 5. IHC • “Immuno” –Refers to antibodies • “Histo” – Refers to tissue • Albert Coons – First implemented (1941) • Principle – Binding of specific antibody to the specific antigen
- 6. Definition •Defined as companiedprocess of anatomical, immunological and biochemical techniques to identify the discrete tissue components by interaction of target antigens with specific antibodies targeted with a visible label.
- 7. Uses 1. Differentiate amonghistologically similar tumour 2. Confirm the histological impression 3. Support the diagnosis of rare tumour 4. Support the diagnosis of unusual location or age
- 8. Protocol Tissue processing Antigen retrieval Inhibitionof endogenous peroxidase Primary antibody incubation Secondary antibody incubation DAB staining Hematoxylin staining Dissolution & Clearing visualization
- 9. Markers for Fibroblastic/Myofibroblastic & Fibrohistiocytictumour 1. Intermediate filament 1. Vimentin 2. Keratin 3. Glial fibrillary acid protein 2. Actin 3. Desmin 4. CD 34 5. CD 68
- 10. Vimentin • Cytoplasmic marker •Expressed In all mesenchymal cells 1. Fibrocyte 2. Fibroblast 3. Smooth muscle 4. Endothelium 5. Schwann cells 6. Renal tubules 7. Thyroid follicular cells
- 11. • Used –Diagnosis of carcinomas of uncertain primary site • Strong expression – Clue to Renal, Endometrial & Thyroid carcinoma Pitfall: • Use of single marker is limited diagnostic value • Because of co expression with other cytokeratins
- 12. Strong positive (>90%) 1.Nodular fasciitis 2. Fibrous histiocytoma 3. Myofibroblastoma 4. Angiomyxoid fibroma 5. Giant cell angiofibroma 6. Dermatomyofibroma 7. Solitary fibrous tumour 8. Congenital & infantile fibro sarcoma 9. DFSP
- 13. Cytokeratin • Cytoplasmic marker •Epithelial cells • Composed of > 20 isotypes • Type I – Acidic – 9 to 20 • Type II – Basic – 1 to 8 • Expressed both carcinomas and sarcomas • Different CK expressed in different epithelial types at different stage of differentiation
- 14. Sarcomas Usual Unusual Synovial sarcomaNeuroendocrine tumour Epithelioid sarcoma Metastatic melanomas Wilm’s tumour Rhabdomyosarcoma Pit falls: Also expressed various non epithelial tissue & tumours
- 15. Actin • Cytoplasmic marker •Seen in pure myogenic differentiation • 6 isoforms • Antibodies used – HHF35,1A4 Pit fall : +ve other than smooth muscle tumour 1. Endometrial stromal tumour 2. Synovial sarcoma 3. GIST’s
- 16. Strong positive 1. Myofibroblastoma 2.Angiomyxoid fibroma 3. Dermatomyofibroma 4. Solitary myofibroma 5. Angioleiomyoma 6. Leiomyoma 7. Leiomyosarcoma
- 17. Desmin • Intermediate filament •Present in cardiac , skeletal muscle & Smooth muscle • Strong positive 1. Myofibroblastoma 2. Angiomyofibroblastoma 3. Solitary myofibroma 4. Rhabdomyoma 5. Rhabdomyosarcoma Pit fall: +ve other than RMS like desmoplastic small round cell tumour
- 18. CD 68 •Expressed onlyin multinucleated cells •+ve: 1. GCT of tendon sheath 2. Tenosynovial giant cell tumour
- 19.
- 20. 1. Actin 2. Desmin 3.Myogenic transcription factors 4. Caldesmon 5. Calponin 6. Myoglobin 7. Myosin
- 21. Myogenic transcription factor •Role – Differentiation of mesenchymal progenitors to myogenic lineage • Myo D1 & Myogenin – expressed only in fetal skeletal tissue • Not in adult skeletal tissue • Only nuclear positivity considered • +ve: • Embryonal & Alveolar RMS Pit falls: Also positive in non neoplastic myoblast,desmoid tumour, Wilm’s tumour
- 22. Caldesmon • Cytoplasmic marker •Calcium & Calmodulin binding protein • Role – Contraction of smooth muscle • 2 isoforms are available ( LMW & HMW) • Expressed in Smooth muscle & Myoepithelial cells • Not seen in myofibroblast • +ve : LM,LMS,GIST’S, Glomus tumour • -ve : RMS Pit fall: +ve in non smooth muscle lesion GIST
- 23. Calponin • Cytoplasmic marker •Actin , Tropomyosin & Calmodulin binding protein • Role – Smooth muscle contraction • Expressed in smooth muscle, myoepithelial cells, myofibroblast • +ve : LM,LMS,NODULAR fasciitis • -ve : GIST’s
- 24. Myoglobin • Cytoplasmic marker •Iron & o2 binding polypeptide • Expressed in Skeletal muscle , Cardiac muscle, Rhabdomyoblast • Lack of expression in Smooth muscle Pit fall : • Also expressed in various other carcinomas ( BREAST,PROSTATE,COLON)
- 25.
- 26. 1. S 100 2.MDM 2 3. CDK 4 4. P16
- 27. S 100 • Calciumbinding protein • Acidic nature • 3 isoforms • Expressed in normal and neoplastic cells
- 28. Normal cell Neoplasticcell Melanocyte Nevi, Malignant melanoma Adipocyte Lipoma , Liposarcoma Schwann cell Schwannoma, Neurofibroma Cartilage Chondroma, chondrosarcoma Langerhans cells Langerhans cell histiocytosis
- 29. MDM 2 • Murinedouble minute 2 • Nuclear & Cytoplasmic maker • + ve in Liposarcoma Pit fall: • Also expressed in many sarcomas
- 30. CDK 4 • Cyclindependent kinase 4 • Nuclear marker • Role – Regulation of cell cycle • Expressed in some Epithelial & Mesenchymal cells Pit fall: • Over expression in Liposarcoma , Osteosarcoma
- 31. P 16 • Cyclindependent kinase inhibitor 2A • Tumour suppressor protein • Expressed in few carcinoma types & HPV associated SCC of different origin • +ve : Well differentiated , Dedifferentiated Liposarcoma • -ve : Normal fatty tissue & Lipoma
- 32.
- 33. 1. CD 31 2.CD 34 3. Factor 8 4. Podoplanin 5. Thrombomodulin CD 141 6. CD 105 7. VEGF-3
- 34. CD 31 • Commonvascular marker • Platelet endothelial cell adhesion molecule 1 • Most sensitive & specific marker • Not seen in non endothelial cells • Expressed • Endothelial cell unction • Platelet,monocyte,granulocyte • +ve : Angiosarcoma, Haemangioendothelioma, Hemangioma,kaposi’s sarcoma
- 35. CD 34 • Hematopoieticprogenitor cell antigen • Expressed in precursor of Myeloid, Lymphoid lineage & Mesenchymal cell • Widely used • But less specific than CD 31 • +ve : • Kaposi’s sarcoma, Angiosarcoma,DFSP, Solitary fibrous tumour, Neurofibroma Pit fall: • Broad spectrum of expression used as screening marker
- 36. Factor 8 • Cytoplasmicmarker • First specific marker for endothelium used in IHC • Less sensitive • +ve : Vascular tumour Pit fall: • Also present in serum • +ve in tumour necrosis & hemorrhage
- 37. Podoplanin • Membranous /cytoplasmic markers • Known as C2-40 • Expressed in • Fetal germ cells • Membrane of several mature cells +ve : Lymphangioma, Mesothelioma, Adenomatoid tumour Pit fall: • Broad spectrum of expression • Used with other specific antibodies
- 38. Thrombomodulin • Known asCD 141 • Expressed in Endothelium, Trophoblast, Mesothelial cells • +ve : • Hemangioma • Haemangioendothelioma • Angiosarcoma • Mesothelioma
- 39. VEGF-3 • Transmembrane receptortyrosine kinase specific protein • Expressed in Endothelial & Trophoblast • +ve : 1. Kaposi’s sarcoma 2. Angiosarcoma
- 40.
- 41. 1. Synaptophysin 2. Chromogranin 3.Neuron specific enolase 4. Neuro filament protein
- 42. Synaptophysin • Membrane channelprotein • Expressed 1. Neural cells 2. Neuroendocrine cell +ve : 1. Neuroblastoma 2. Ganglioneuroma 3. Medulloblastoma 4. carcinoids
- 43. Chromogranin • Calcium bindingprotein • Expressed • Neural cells • Neuroendocrine cells • +ve : • Paragangliomas • Pheochromocytoma • Carcinoids • -ve : • PNET
- 44. Neuron specific enolase •Expressed in neural & neuroendocrine cells • +ve: 1. Paragangliomas 2. Carcinoids 3. Melanomas 4. Neuroblastoma 5. Leiomyosarcoma 6. Angiosarcoma
- 45. Neuro filament proteins •Intermediate filaments of Neurons & Axons • 3 types • +ve : 1. Neuroblastoma 2. Merkel cell carcinoma 3. Adrenal Pheochromocytoma
- 46.
- 47. CD 99 CD56 Transmembrane glycoprotein Neural cell adhesive molecules +ve Ewing’s PNET Lymphoma +ve Synovial sarcoma MPNST Schwannoma RMS,LMS -ve -ve
- 48. Osseous & Cartilaginous marker 1.S 100 2. Ostocalnin 3. Ostonectin 4. Androgen receptor 5. Pan cytokeratin
- 49. S 100 Ostonectin Softtissue Chondroma Chondroblastoma Mesenchymal osteosarcoma Extra skeletal osteosarcoma
- 50. Melanocytic markers 1. HMB-45 2.Throsinase 3. Melan –A 4. Microphthalmic transcription factor
- 51. HMB 45 • Organellespecific marker • Detect immature melanocytes • +ve: 1. Malignant melanoma 2. Angiosarcoma 3. Lymphomatosis • -ve: 1. Intradermal nevus
- 52. Thyrosinase • Cytoplasmic marker •Catalyze tyrosine – Incorporation into the melanin pigment • Excellent for Malignant Melanoma
- 53. Melan -A • Markersof melanosomes • Not for melanocytes • +ve : 1. Epithelioid melanomas 2. PECOMA’s 3. Angiomyolipoma
- 54. Microphthalmic transcription factor • Nuclearmarker • +ve: 1. Primary melanomas 2. Metastatic melanomas 3. Clear cell sarcomas 4. Leiomyosarcoma
- 55. Prognostic markers 1. Ki67 & analogs - Indicate distant metastasis 2. P 53 - High grade tumour - Worst outcome 3. P16 & P27
- 56. Cell type TumourDiagnosis Round cell Rhabdomyosarcoma PNET Myo D1 CD 99 Spindle cell Schwannoma Neurofibroma Kaposi’s sarcoma DFSP MPNST S -100 S – 100 CD 34/CD 31 CD 34 S - 100 Epithelioid cell Angiosarcoma Clear cell sarcoma Epithelioid sarcoma Angiosarcoma CD 31/34 Melanoma markers CD 34 CD 31/34
- 57. Antibody Significance Actin Smoothmuscle cell CD 31 Endothelial cells CD 34 Precursor of blood & Endothelial cell Cytokeratin Epithelial origin Desmin General marker for muscle cells HMB 45 Melanosomes Neurofilament Nervous tissue Neuron specific enolase Neuroendocrine cells S 100 Adipose cell, nervous cell, macrophage Vimentin Connective tissue
- 58. Sarcomas IHC Smooth ,Skeletal, Endometrial stromal Desmin Vascular sarcomas CD 31/34 Sarcomas with NEURAL,LIPOMATOUS differentiation S 100 Synovial sarcomas Cytokeratin, CD 99 GIST CD 117 Chondro sarcoma CD57, Vimentin Ewing’s/ PNET CD 99, S100 Desmoplastic round cell tumour Cytokeratin , CD 99, Desmin Osteosarcoma CD 57, Vimentin, EMA