Efficient immunohistochemical differential diagnosis of undifferentiated neoplasia

Conventional histomorpholgy: pleomorph and/or high-grade spindle-cell Conventional histomorpholgy: epitheliod, intermediate-cell to large-cell Conventional histomorpholgy: small-cell

Localisation of tumour Localisation of tumour Localisation of tumour Antibody panel Marker constellation and valuation Antibody panel Marker constellation and valuation Antibody panel Marker constellation and valuation infiltrates infiltrates infiltrates

CK8/18 + (diffuse), Ber-EP4 +/-, GFAP -/+ (focal) p63 +, WT1 -, D2-40 (Podoplanin) -/+ carcinoma metastasis if plexus carcinoma is excluded pleura carcinosis due to non-keratinising squamous cell carcinoma CK8/18, Ber-EP4, GFAP, CK8/18 + (diffuse), Synaptophysin +, TTF-1 +/- Central nervous system GFAP +, CK8/18 -/+ (focal), CD34-posivite microvascular proliferates WT1 + and/or D2-40 (Podoplanin) +, p63 -, Pan-CK + Melan A, CD34 metastasis of a small-cell carcinoma glioblastoma epithelioid or biphasic mesothelioma, Central nervous system CK8/18, GFAP, Melan A, p63, WT1, Podoplanin Attention: Pleura carcinosis due to serous ovarian carcinoma TTF-1, Synaptophysin GFAP +, CK8/18 -/+ (focal), Synaptophysin - Melan A + malignant melanoma (metastasis) Pleura small-cell fraction of a glioblastoma (D2-40), S100, Pan-CK Pan-CK +, other markers negative Pleura carcinosis due to macrocellular carcinoma Melan A + small-cell malignant melanoma (metastasis) Pan-CK + (diffuse or focal, other markers negative) S100 + (diffuse nuclear and cytoplasmic), Pan-CK -/+ (focal) carcinoma metastasis metastasis of a malignant melanoma Pan-CK + (diffuse), Desmin -, CK20 -, Synaptophysin +/- S100 + (diffuse nuclear and cytoplasmic) small-cell carcinoma (if Synaptophysin –: differential diagnosis small- metastasis of malignant melanoma cell squamous cell carcinoma) Pan-CK, S100, CD30, kappa/lambda +, CD20 - immature (anaplastic) plasmocytoma (myeloma) Ber-EP4 + or Ber-EP4 -, Pan-CK +, Calretinin - peritoneal carcinosis Lymph node CK20 + (punctual perinuclear), Synaptophysin + CD20, kappa/lambda Calretinin +, Pan-CK +, Ber-EP4 - merkel cell carcinoma CD30 + (membranous diffuse and in the Golgi region), epithelioid or biphasic mesothelioma, Pan-CK -/+ (focal), CD20 - anaplastic large-cell lymphoma (ALCL) Ber-EP4, Pan-CK, CD99 + (membranous diffuse), Pan-CK -/+ (focal), Synaptophysin +/-, S100 + (diffuse nuclear and cytoplasmic), Pan-CK -/+ (focal), Peritoneum Calretinin, S100, TdT -, S100 -/+ (focal), MPO - PNET CD20 +, CD30 +/- Ber-EP4 -, Inhibin - Pan-CK, S100, CD99, HMB45, Inhibin α α diffuse large-cell B-cell lymphoma, anaplastic variation metastasis of a malignant melanoma other localisation Desmin, Synaptophysin, Synaptophysin +, CD99 -, Pan-CK -, S100-positive sustentacular cells TdT, Myeloperoxidase olfactory neuroblastoma? (rhinopharynx) all markers negative Inhibin α +, Calretinin +, Ber-EP4 -, S100 –/+ (focal) granulosa-cell tumour? carcinoma with lost CK expression or histiocytic sarcoma? (MPO), CK20 S100 + (diffuse nuclear and cytoplasmic) HMB45 +, S100 - perivascular epithelioid-cell tumour small-cell malignant melanoma

Pan-CK + (diffuse or focal, other markers negative) Melan A + small-cell malignant melanoma (metastasis) (sarcomatoid) carcinoma CK8/18 + (diffuse), GFAP -/+ (focal) TdT +, CD99 +/-, Pan-CK -, MPO - S100 + (diffuse nuclear and cytoplasmic), HMB45 +/- carcinoma metastasis after elimination of a plexus carcinoma desmoplastic small- and round-cell tumour? malignant melanoma GFAP +, CK8/18 -/+ (focal) anaplastic glioma or glioblastoma MPO + myeloid sarcoma Gastrointestinal tract, Central nervous system CK8/18, GFAP, Melan A, genito-urinary system, Pan-CK, S100, CD30 + (membranous diffuse and in the Golgi region), OCT4, CD117 Melan A + malignant melanoma (metastasis) parenchymatous organs, HMB45, CD30, CD20 Pan-CK -/+ (focal), CD20 - anaplastic large cell lymphoma (ALCL) OCT4 +, CD117 - embryonary carcinoma head/neck area CD20 +, CD30 +/- OCT4 +, CD117 - germinoma diffuse large-cell B-cell lymphoma, anaplastic variation Marker for immunohistochemical diagnostics of CUP (Carcinoma of Unknown Primary) all markers negative Marker Predicted localisation of Type of carcinoma Comments carcinoma with loss of CK expression or histiocytic sarcoma? Pan-CK + (diffuse), OCT4 - carcinoma metastasis primary tumour AFP liver HCC Positive in yolk sack tumours S100 + (diffuse nuclear and cytoplasmic), Pan-CK -/+ (focal) metastasis of a malignant melanoma Adenocarcinomas of stomach, oesophagus, pancre- Pan-CK+/-, CK5/6 +/-, CD31 -, h-Caldesmon -, S100 -/+ (focal) adenocarcinomas (ovary only in CDX-2 (a) colorectum, ovary, bladder case of gastrointestinal mucinous as, and biliary ducts frequently show heterogeneous (sarcomatoid) carcinoma, differential diagnosis metastasis OCT4 +, Pan-CK + (diffuse), CD30 + (diffuse immunoreactivity) immunoreactivity. Adenocarcinomas of the lung are Pan-CK, CD20, S100, metastasis of an embryonary carcinoma differentiation) rarely positive. S100 + (diffuse nuclear and cytoplasmic) Lymph node OCT4, CD30 OCT4 +, Pan-CK -/+ (focal), CD30 - metastasis of a seminoma (spindle-cell) malignant melanoma CDX-2 (b) middle intestine well-differentiated neuroendocrine CD30 + (membranous diffuse and in the Golgi region), Pan-CK -/+ (focal), CD20 (appendix, ileum) tumours Pan-CK, CK5/6, CD31, CD31 + angiosarcoma Skin anaplastic large-cell lymphoma, „small-cell” variant CK7/CK20 colorectum adenocarcinomas (CK7-/CK20+) S100, CD30, h-Caldesmon h-Caldesmon + (more than single cells) leiomyosarcoma CD20 + diffuse large-cell B-NHL DPC4/SMAD4 pancreas, biliary ducts adenocarcinomas Only the loss of expression is relevant for diagnosis! CD30 + (membranous diffuse and in the Golgi region), Pan-CK -/+ (focal) (sarcomatoid) anaplastic large-cell lymphoma Rare immunoreactivity in adenocarcinomas of the GCDFP-15 breast, skin adnex adenocarcinomas lung. all markers negative Pan-CK +, EMA +/-, Inhibin α -, OCT4 - atypical fibroxanthoma (Head/Neck, actinic lesion?) carcinoma (primary tumour or metastasis) Positive in malignant melanomas and in yolk sack Glypican-3 liver HCC (Hepato Cellular Carcinoma) tumours; otherwise more sensitive and specific than S100 + (diffuse nuclear and cytoplasmic), Pan-CK -/+ (focal), Inhibin α - HepPar1 and TTF1 (cytoplasmic) malignant melanoma Positive in appr. 50% of all adenocarcinomas of S100 + (diffuse nuclear and cytoplasmic), HMB45 + Pan-CK, EMA, S100, Ovary Inhibin α +, EMA -, S100 -/+ (focal) germ band or stroma tumour stomach and to a lesser degree in other primary lo- malignant melanoma, differential diagnosis clear cell sarcoma (if HMB45 –: Inhibin α, OCT4, CD20 HepPar1 liver HCC (Hepato Cellular Carcinoma) calisations (colon, lung, pancreas); therefore minor differential diagnosis malignant peripheral nerve sheath tumour, MPNST) OCT4 +, Pan-CK -/+ (focal) embryonary carcinoma positive predictive value for differentiation from CD31 + angiosarcoma OCT4 +, Pan-CK -/+ (focal) dysgerminoma HCC and liver metastases. Myf4 + (nuclear, attention: immunoreactivity in regenerative skeletal CD20 + diffuse large-cell B-NHL Estrogen Receptor Mamma, ovary, corpus uteri adenocarcinomas muscle fibres in tumour-infiltrated areas) Parathyroid hormone parathyroid gland adenomas and carcinomas S100, HMB45, Myf4, clear-cell and papillary kidney-cell Soft tissue pleomorphic rhabdomyosarcoma or other tumour with focal rhabdo- PAX-2 kidney, ovary Mesotheliomas and HCC negative CD31, h-Caldesmon, CD30 myoblastic differentiation Pan-CK + (diffuse) carcinoma carcinoma, serous ovary carcinoma h-Caldesmon + (more than single cells) S100 + (diffuse nuclear and cytoplasmic), Pan-CK -/+ (focal), Inhibin α - PSA prostate adenocarcinomas pleomorphic leiomyosarcoma malignant melanoma, (if HMB45 –: differential diagnosis malignant Gastrointestinal tract, Immunoreactivity in appr. 10-30% of mesothelio- peripheral nerve sheath tumour, MPNST) CD30 + (membranous diffuse and in the Golgi region) Respiratory tract, mas, breast carcinomas, prostate carcinomas, ovary Pan-CK, EMA, S100, (sarcomatoid) anaplastic large-cell lymphoma genito-urinary system, HMB45 +, S100 - perivascular epithelioid-cell tumour RCC kidney carcinomas carcinomas, lung carcinomas, and adrenocortical Inhibin α, OCT4, CD20 neoplasias. Adenomas of the parathyroid gland are other marker combination parenchymatous organs, CD20 + diffuse large-cell B-NHL 100% positive; clear-cell tumours of the skin with sarcoma; needs further investigation head/neck area CD30 + (membranous diffuse and in the Golgi region), Pan-CK -/+ (focal), CD20- varying histogenesis are negative. anaplastic large-cell lymphoma, „small-cell” variant Thyreoglobulin thyroid gland differentiated and insular thyroid Kappa/lambda +, CD20 - gland carcinomas plasmoblastic lymphoma? (HIV + ?), ALK-1-positive diffuse large-cell adenocarcinomas, large-cell non- B-NHL? TTF1 (a) nuclear neuroendocrine carcinomas; not (only clone 8G7G3/1, lung applicable for small-cell carcinomas With clone 8G7G3/1 also rare immunoreactivity in other clones mostly (expression not localisation-specific) endometrial adenocarcinomas. less specific) and squamous cell carcinomas (no Pan-CK + (diffuse) epithelioid sarcoma, differential diagnosis expression) carcinoma metastasis TTF1 (b) nuclear lung carcinoid S100 + (diffuse nuclear and cytoplasmic), HMB45 +/- malignant melanoma, differential diagnosis clear-cell sarcoma differentiated and insular carcino- Pan-CK, S100, CD31, TTF1 (c) nuclear thyroid gland Soft tissue (if HMB45 –: differential diagnosis malignant peripheral nerve sheath mas HMB45, CD117 TTF1 cytoplasmic tumour, MPNST) liver HCC (Hepato Cellular Carcinoma) Similar sensitivity but slightly more specific than HMB45 +, S100 - perivascular epithelioid-cell tumour (only clone 8G7G3/1) HepPar1 CD31 + epithelioid angiosarcoma Uroplakin III urinary tract urothelial carcinoma Brenner tumours of the ovary are positive CD117 + epithelioid gastro-intestinal stroma tumour (GIST)? WT1 (nuclear) ovary serous adenocarcinoma Mesotheliomas are also positive

This IHC algorithm was prepared to the best of our knowledge. However, in special cases (for example childhood tumours) the information can be incomplete and the user has to be aware of this. The use of this diagram is solely the responsibility of the user. CDX-2, liver metastasis Estrogen Receptor, liver metastasis of HepPar1, Under no circumstances shall Zytomed System be liable for any damages arising out of the use of this table. of colon carcinoma breast carcinoma hepatocellular carcinoma We kindly would like to thank Dr. med. habil. Olaf Kaufmann for his expert advice in the preparation of this diagram. © Zytomed Systems 2009

Synaptophysin, CDX-2, Cytokeratin 20, Oct-4, seminoma Calretinin, mesothelioma TTF-1 (clone 8G7G3/1), normal liver tissue neuroendocrine tumour colon carcinoma colon carcinoma