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What are triple negative breast cancers? A synopsis of their histological patterns Ian Ellis Molecular Medical Sciences, University of Nottingham Department of Histopathology, Nottingham University Hospitals NHS Trust

ER neg ER pos Sorlie, et al. PNAS 2001

Two main groups: luminal (majority) and basal groups The concept of BP has been known for some time First described using electron microscopy >30 years age Its potential poor survival first reported by Dairkee et al in 1987 High frequency in BRCA1 gene mutation carriers

Additional molecular subtypes enriched for TNBCs: claudin-low - EMT phenotype molecular apocrine - activated AR pathway. basal-like 1 basal-like 2 luminal AR immunomodulatory mesenchymal mesenchymal stem-like Basal like/immune-suppressed basal-like/immune activated luminal AR mesenchymal

Special types of breast cancer are more homogeneous at the transcriptome level Luminal Basal-like Mol apocrine Mucinous A Classic ILC/ Tubular Mucinous B Neuroendocrine Micropapillary Adenoid cystic Medullary Metaplastic Pleomorphic ILC Apocrine Weigelt et al. J Pathol 2008

Triple Negative Breast Cancer

HER 2

Ck 5 6

Ck 14

Association between BP and survival, whole series

Overlap of BRCA1 and Basal-like Genotypes Basal Sub-Type BRCA1 mutated Sorlie PNAS 2003

Familial Breast Cancer BRCA 1 Phenotype Histological features High grade and mitotic count Medullary or atypical medullary type Molecular features ER & PR negative HER2 negative P53 positive Basal ck positive

Familial Breast Cancer BRCA 1 basal phenotype Ck 5/6&14 +ve - 44% of all BRCA 1 carriers Ck 5/6&14 +ve < 2% sporadic cancers Lakhani Clin Cancer Res 2005 11 5175

Basal-like carcinomas BRCA1 tumours Ploidy Aneuploid Aneuploid Most frequent copy number gains Most frequent copy number losses 1q 3q 6p 7p 7q 8q 10p 17q22-qtel 21q 3p 4q 5q 6q 12q 13q 14q 1q 3q 7p 8q 10p 12p 17q22-qtel 3p 4p 5q 12q 16p 18q BRCA1 gene mutations Absent Present

BRCA1 downregulation High histological grade Medullary histological type Basal-like immunophenotype

Hypothesis BRCA1 inactivation in Basal-Like cancers Gene promoter methylation Transcriptional inactivation

BRCA1 methylation ER ve PR ve High histological grade Medullary histological type Catteau et al. Oncogene, 1999; Esteller et al. JNCI 2000; Osin et al. Int J Surg Pathol, 2003

BRCA1 methylation in metaplastic breast carcinomas U M U M U M U M U M U M U M U M BRCA1 gene promoter methylation 17/ 27 (63%) Type BRCA1 M BRCA1 U Metaplastic 17 10 IDC-basal 4 25 p<0.0005

Basal-like carcinomas? BRCA1 downregulation BRCA1 methylation Salivary gland-like tumours IDC Basal-like Medullary Metaplastic

WHO 2012 Carcinoma with medullary features Definition - Carcinomas with medullary features include medullary carcinomas (MC) atypical MC a subset of invasive ductal carcinomas of no special type These tumours demonstrate all or some of the following features: a circumscribed or pushing border a syncytial growth pattern cells with high-grade nuclei prominent lymphoid infiltration.

Metaplastic ( sarcomatoid ) Carcinoma Histological classification Monophasic sarcomatoid carcinoma or spindle cell carcinoma Biphasic sarcomatoid carcinoma or malignant mixed tumour (carcinosarcoma) Low grade adenosquamous carcinoma

Metaplastic ( sarcomatoid ) Carcinoma Microscopic features Monophasic sarcomatoid carcinoma Composed predominantly of elongated plump cells of relatively monotonous appearance Mitoses are variable in number but are usually plentiful An invasive epithelial element or foci of high grade DCIS can be identified in a minority of cases

Metaplastic ( sarcomatoid ) Carcinoma Biphasic sarcomatoid carcinoma Microscopic features Mesenchymal element is usually fibrosarcomatous or malignant fibro-histiocytoma-like More rarely chondrosarcomatous, osteosarcomatous, leiomyosarcomatous or angiosarcomatous elements are present The epithelial component is usually of ductal NST pattern, grade 2 or 3 but may have squamous features High grade DC1S is present in up to 50% of cases

Low grade adenosquamous carcinoma Originally described by Rosen in 1987, Am J Surg Pat Lesions characterised by small glandular structures with varying amounts of epidermoid differentiation embedded in a collagenous stroma Tendency to grow between and around ducts and lobules Round to oval glands Can have spindle cell and syringomatous areas Squamous component 5-80% Clusters of inflammatory cells at the periphery All ER/PgR negative

Low grade adenosquamous carcinoma Association with underlying fibrosclerotic lesions including complex sclerosing lesions, papillary lesions and nipple adenomas Lymph node involvement rare Metastatic disease - exceptional

Triple Negative Breast Cancer

Salivary gland like tumours of the breast Benign Mixed tumour Adenomyoepithelioma Benign myoepithelioma Malignant Acinic cell carcinoma Adenoid cystic carcinoma Low grade adenosquamous carcinoma Oncocytic carcinoma Mucoepidermoid carcinoma Malignant myoepithelioma

Adenoid cystic carcinoma 0.1 1% of breast cancers Wide age range Mass lesion, often periareolar, may be painful Well defined Excellent long term prognosis ->90-100% 10 year survival (cf salivary gland tumours)

Epithelial cells EMA CD117 CK7

Basaloid cells p63 vimentin CK14

Adenoid cystic carcinoma Cribriform, solid, tubular, reticular, basaloid patterns Dual population: epithelial and basaloid cells Epithelial: CK7, CEA, EMA, CD117 Basaloid: CK14, CD17, vimentin,s100, actin, calponin, p63 May be associated with microglandular adenosis

Adenoid cystic carcinoma- molecular pathology Clusters with metaplastic and medullary carcinomas triple negative Translocation t (6;9) (q22-23; p23-24) similar to salivary and other adenoid cystic carcinomas

AcCC of the breast Clinicopathological features First report by Roncaroli 1996 Acinic cell-like carcinoma of the breast. Virchows Arc 1996;429:69-74 Defined by serous differentiation Infiltrative margin Microglandular areas merging with solid aggrageates Intraluminal inspisssated secretion Stroma can be fibro-fatty without desmoplasia

AcCC of the breast Morphology II Abundant eosinophilic or amphophilic granular cytoplasm Variable mitotic count Immunohistochemistry: + Luminal cytokeratins, S100, Lysosyme, alfa 1- antitrypsin, alfa-amylase(focal, scanty), IgA, E Cadherin - Basal cytokeratins, ER, PR, HER 2, GCDFP EM: membrane bound zymogen granules 0.08-0.9um

Actin

Actin Amylase

Actin Lysozyme

S100 Actin

Actin

Differential diagnosis Tumours with granular cytoplasm Neuroendocrine carcinoma ICH Oncocytic tumours mitochondria Apocrine carcinoma GCDFP 15 Tumours with architectural similarities Microglandular adenosis Secretory carcinoma

AcCC v MGA Overlapping arhitectural features-small glandular/acinar structure without myoepthelial layer IHC similarities- CK,S100,Lysosyme, ER, Her2 Morphological similarities between MGA and AcCC Transitional forms do exist Reported carcinomas associated with MGA retain the acinar architecture High rate of invasive carcinoma reported with MGA But AcCC usually with solid areas Lack of BM IHC differencies-ema,?amylase Zymogen granules on EM

AcCC v. Secretory carcinoma Secretory carcinoma is analogous to AcCC of microcystic, follicular and papillary cystic type Similar IHC:Lysosyme+, S100+, alfa 1-antitrypsin+, alfa-amylase+, IgA+ ER-, Her 2- Hirokawa Histopathology 2002, 40, 223-29

Secretory carcinoma First identified by McDivitt & Stewart in 1966 as a children s breast cancer but later recognised as also occuring in young, and a few adults. Male association Usually present as a well defined sub-areolar mass Prognosis is favourable and is thought to be better in children than in adults Local recurrences, if developed, are late Lymph node metastasis are uncommon Distant metastasis are exceedingly rare Death is unusual, but has been reported

D PAS

Secretory carcinoma Low nuclear grade with vacuolated cytoplasm which may contain eosinophilic secretion arranged in cribriform patterns with the spaces containing eosinophilic secretions Typically, they show strong reactivity with S100 They are mostly triple negative Express basal cytokeratins, and belong to the basallike molecular group of breast cancers Genetically they are characterised by the presence of a chromosomal translocation t(12;15)(p13;q25) which results in the formation of ETV6-NTRK3 fusion gene

Triple Negative Breast Cancer

Do not forget

Invasive Apocrine Carcinoma Rare subtype in pure form 0.3 4.0% Focal apocrine differentiation common 60% NST on morphology 72% express GCDFP Lack of robust criteria

Invasive Apocrine Carcinoma Presentation, prognostic characteristics and behaviour similar to NST Immunophenotype ER neg PR neg AR pos

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