5/25/2017 The Double-Edged Sword of Immunostains The Double-Edged Sword of Immunostains in Diagnosis of Breast Pathology in Diagnostic Breast Pathology Diagnostic Help Yunn-Yi Chen, MD, PhD Professor Director of Immunohistochemistry Laboratory UCSF Diagnostic Pitfall Use of IHC in Diagnosis of Breast Pathology � Distinction of noninvasive from invasive lesions You mean you want to talk about � Measurement of biomarkers why IHC will kill you both ways? � Assessment of ductal proliferative and papillary lesions � Differentiation between ductal and lobular CIS ? � Workup of spindle cell lesions � Diagnosis of metastatic tumors in the breast � Evaluation of sentinel lymph nodes 1
5/25/2017 Markers Staining Myoepithtelial Cells (MEC) Markers staining myoepithtelial cells (MEC) Cytoplasmic(+ membranous) Cytoplasmic/nuclear Nuclear Cytoplasmic(+ membranous) Cytoplasmic/nuclear Nuclear SMA Calponin S100 p63 SMA Calponin S100 p63 SMM basal CKs SMM CK5/6 CD10 D2-40 CD10 D2-40 h-caldesmon P-cadherin h-caldesmon P-cadherin GFAP WT1 GFAP WT1 Maspin Nestin Maspin Nestin p75 CD109 p75 CD109 Panel of at least two markers-- Stratifin CD44s Stratifin CD44s Muscle-specific actin Muscle-specific actin p63 + cytoplasmic marker (SMM or calponin) Caveolin 1 and 2 Caveolin 1 and 2 Metallothionein Metallothionein …… …… Pitfalls in Interpreting MEC Markers Comparison of Reactivity by MEC Markers Marker Myoepi. Myofibro- Vessels Carcinoma � Stromal (myofibroblast and vessel) staining cells blasts cells SMA +++++ +++ +++ Rare + � Tumor cell staining Calponin ++++ to +++++ ++ +++ Rare + � Biology of the lesions SMMHC ++++ + +++ Rare + (SMM) - p63 ++++ - Occasional + � Artifact in interpretation (15.7 to 23% IDC)* CK5/6 (other +++ to ++++ - - Occasional + HMW CK) (~10% IDC)¥ *p63 positivity in ~100% adenoid cystic carcinoma and majority of metaplastic carcinoma ¥CK5/6 positivity more likely to be seen in high grade IDC and DCIS 2
5/25/2017 Myofibroblast Staining Mimicking ME Cells Myofibroblast Staining Mimicking ME Cells calponin calponin p63 SMA may be helpful in suboptimally-fixed tissue Myofibroblast Staining Mimicking ME Cells calponin SMA > calponin > SMM � Not seen with p63 or CK5/6 � calponin SMA p63 SMM 3
5/25/2017 Myofibroblast Staining Mimicking ME Cells Myofibroblasts around inv. gland ME cells around DCIS Myofibroblasts around inv. gland ME cells around DCIS SMM SMM stain p63 stain Tumor Cell Staining from MEC Markers Pitfall of Tumor Cell Staining-- � Location and shape of positive nuclei More common with p63 and � � Intensity of staining CK5/6 Rarely with SMM and � calponin p63 p63 SMM 4
5/25/2017 Pitfall of Tumor Cell Staining-- Pitfalls in Interpreting MEC Markers � Stromal (myofibroblast and vessel) staining � Tumor cell staining � Biology of the lesions p63 SMM � Phenotypic alterations in DCIS-associated ME cells � Phenotypic alterations in ME cells-associated with benign sclerosing lesions � Non-invasive lesions without expression of MEC markers � Invasive carcinomas with expression of MEC markers � Artefact in interpretation Reduced MEC Marker Expression in DCIS Phenotypic Alterations in DCIS-associated ME Cells SMM � Reduced expression to focal absence of one or more MEC markers in DCIS-associated ME cells � Incidence of attenuated expression � Overall: SMM (77%) > CK5/6 (30%) > calponin (17%) > p63 (13%) > SMA (1%) p63 SMA � However, variable in each case Hilson et al: Am J Surg Pathol 2009 5
5/25/2017 Papillary DCIS often with attenuated MEC expression around the ducts Round cribriform glands, negative p63, SMM and calponin Attenuated MEC staining in cribriform DCIS or invasive cancer? SMM p63 Cribriform DCIS or Invasive Cribriform carcinoma? Phenotypic Alterations in ME Cells Associated with Benign Sclerosing Lesions of the Breast � Reduced expression to focal absence of one or more MEC markers in ME cells associated with benign sclerosing lesions � Incidence of attenuated expression � Overall: CK5/6 (32%) > SMM (21%) > p63 (9%) > calponin (6%) > SMA (0%) � However, variable in each case Hilson et al: Am J Surg Pathol 2010 6
5/25/2017 Patchy Attenuated MEC Staining in RSL Reduced MEC Marker Expression in Radial Sclerosing Lesion Almost complete absence of staining for p63 p63 p63 SMM Variably Reduced MEC Marker Expression in Radial Sclerosing Lesion p63 Microglandular Adenosis (MGA)-- A Noninvasive Glandular Lesion Without Expression of MEC Markers SMM CK5/6 7
5/25/2017 Microglandular Adenosis-- Microglandular Adenosis-- Hypocellular collagenous stroma Haphazard distribution Microglandular Adenosis Microglandular Adenosis-- SMM Uniform small glands, open lumen, eosinophilic secretion PAS stain p63 ER 8
5/25/2017 MGA--Pitfall in Interpreting MEC Markers Microglandular Adenosis � Red flag: ER/PR negative “well-differentiated invasive ER S100 ductal carcinoma” � Characteristic H&E morphologic features Uniform small round glands with open lumen and PAS+ � eosinophilic secretion Hypocellular collagenous/fatty stroma � � S100 diffusely and strongly + Low-grade Adenosquamous Carcinoma (LGASC) Invasive Carcinomas Expressing MEC Markers-- Pitfall in Interpreting MEC Markers � Carcinomas with myoepithelial differentiation � Adenoid cystic carcinoma (AdCC) � Low grade adenosquamous carcinoma (LGASC) � Neoplastic MEC: Variable and patchy expression of individual MEC markers, typically p63 positive � Misleading peripheral staining (esp. p63) � Patchy and variable staining (SMM, calponin) � Multi-layering of MEC marker-positive cells (p63) 9
5/25/2017 LGASC-- Peripheral Staining for p63 Low-grade adenosquamous carcinoma-- p63 Variable expression of MEC markers (positive p63, negative SMM) p63 SMM Low-grade Adenosquamous CA (LGASC) Low-grade Adenosquamous Carcinoma (LGASC)-- Invasive Carcinoma with positive MEC Markers � p63 positive and variable expression for SMM, calponin Patchy MEC marker expression Multi-layering of p63 positive cells p63 � Characteristic morphologic features Calponin Infiltrative � Spindle cellular stroma, prominent lymphoid reaction � Glands (long, irregular) and solid squamous nests (comma � shaped extension), ± squamous cysts Bland cytology � � ER/PR/HER2 triple negative 10
5/25/2017 Tubular AdCC-- Biphasic Epi-Myoepithelial Diff. Adenoid Cystic Carcinoma (AdCC) May mimic IDC or benign sclerosing lesion � Architectural patterns Cribriform, tubular/trabecular, solid; solid basaloid variant � � Dual epithelial and myoepithelial cell types � ER/PR/HER2 triple negative � t(6;9) MYB-NFIB or t(8;9) MYBL1-NFIB translocation MYB overexpression in 80 to 100% AdCC � � DDx depending on the growth patterns Tubular pattern: mimic benign sclerosing lesion, well-diff. IDC � Myoepithelial type cells: variable expression of MEC markers, � usually p63 +, SMA +/-, and SMM/calponin -/+ Myoepithelial differentiation: pitfall in interpretation of MEC � markers AdCC-- Variable MEC expression and negative ER AdCC-- Variable MEC expression and positive MYB p63 SMM p63 SMM Calponin Calponin ER MYB 11
5/25/2017 MYB IHC as a diagnostic adjunct in AdCC FISH MYB break apart probe � Tests based on MYB-NFIB translocation � FISH: MYB rearrangement Epithelial Displacement-- 50% to 90% � � MYB IHC: diffuse, moderate to Pitfall in Using MEC Markers strong nuclear expression � 80 to 100% MYB � IHC more sensitive and specific assay than FISH for dx of AdCC (Poling et al: Am J Surg Pathol 2017) Epithelial Displacement after Prior Needle Biopsy 63 y F with a left breast mass who underwent a core biopsy followed by excision Breast triple stain 12
5/25/2017 Epithelial Displacement after Prior Needle Biopsy Use of IHC in Diagnosis of Breast Pathology � Common with papillary lesions � IHC often misleading � Distinction of noninvasive from invasive lesions � H&E morphology most helpful Within biopsy tracts � � Measurement of biomarkers Associated granulation tissue, � foamy macrophages, hemosiderin Linear arrangement of glands/nests � � Assessment of ductal proliferative and papillary lesions � Differentiation between ductal and lobular CIS � Workup of spindle cell lesions � Diagnosis of metastatic tumors in the breast � Evaluation of sentinel lymph nodes Breast triple stain Papillary Lesions: Challenging Morphologic Spectrum Papillary Lesions of the Breast (WHO 2012) � Intraductal papilloma Papilloma � with various benign alterations � with ADH involving papilloma (atypical papilloma) � with DCIS involving papilloma (DCIS arising in a papilloma) � Intraductal papillary carcinoma (Papillary DCIS) � Encapsulated (intracystic) papillary carcinoma � Solid papillary carcinoma Papillary CA � Invasive papillary carcinoma 13
5/25/2017 Benign papilloma retains a continuous layer of ME cells along the fibrovascular cores P63 stain MEC markers, CK5/6 and ER-- IHC markers useful in distinguishing papilloma from papillary carcinoma Papillary carcinoma lacks ME cells along the fibrovascular cores Benign Papilloma-- P63 stain � CK5/6 positive � ER patchy and variable CK5/6 ER 14
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