3/29/2017

Disclosure of Relevant What I Learned from 3 Financial Relationships

USCAP requires that all faculty in a position to Cases and 3 Antibodies influence or control the content of CME disclose any relevant financial relationship WITH COMMERCIAL INTERESTS which they or their Melinda Sanders, M.D spouse/partner have, or have had, within the past 12 months, which relates to the content of this educational activity and creates a conflict of interest. Vanderbilt University Medical Center Dr. Melinda Sanders declares he/she has no conflict(s) of interest Professor of Pathology to disclose. Consultant in Breast Pathology

Case #1 History Left Breast Biopsy

• 61 yo female with stage 4 appendiceal adenocarcinoma • 2.1 cm left breast mass and an enlarged axillary lymph node found during staging work‐up • Core needle biopsy performed

Left Breast Biopsy

Left breast biopsy

1 3/29/2017

Appendiceal adenocarcinoma

Breast biopsy

Biopsy #1 Immunohistochemistry Repeat Needle Core Biopsy of Breast

GATA3 ‐2 ‐2 SOX10

Negative expression‐ GATA3, ER, PR, CK7, CK20, CDX‐2 Diagnosis: Poorly differentiated malignant neoplasm and skeletal muscle, cytologically different from abdominal mutinous adenocarcinoma (CDX‐2+) Note: Does not exclude a breast primary

2 3/29/2017

Biopsy #2 Immunohistochemistry Diagnosis ‐ Malignant CK AE1/3 GATA3 CK 7CK 8/18 • Wire‐localized partial mastectomy‐ • 17 mm Malignant Melanoma • SNL –1/1 lymph node S100 SOX10 MITF MelanA HMB 45 • 2.5 mm metastasis • LN dissection‐ • 0/19 lymph nodes

Case #1 Melanoma is most common Keratin Expression in Melanoma Melanoma non‐hematopoietic malig to met to AE 1/3 + patchy weak‐mod breast, may rare occur as primary CK7 ‐ • Keratin expression typically helps support a diagnosis of carcinoma and Breast ER ‐ Think about melanoma when … “rules out sarcoma, lymphoma, and melanoma” GATA3 ‐ • Expression not infrequent‐ most common AE 1/3, CK8, CK18, MNF‐116 • ER‐/PR‐ tumor, nested CK 8/18 ‐ • Aberrant keratin expression more common in metastatic than primary S100 + morphology, especially if TILs • 4‐10% primary SOX10 + • Aberrant CK expression = major • 40‐70% metastatic melanoma Melanoma MITF + pitfall, especially in melanoma • Implies a correlation between keratin expression and a more aggressive MelanA + metastases. tumor phenotype. Safadi et al.,Oral and Maxillofacial Pathol 2016;121:510‐519 HMB45 + • If GATA3 negative, perform Plaza et al., App Immunohistochem Mol Morphol 2007;15:421‐425 GI CDX2 ‐ panel of MM markers HER2 ND

Percentage of Primary vs. Metastatic Melanoma Cases with Keratin Expression S‐100 is Highly Sensitive but not Specific for MM MNF CK18 116 • Normally found in cells derived for including CK8 and myoepithelial cells • Most sensitive melanoma marker (>95%), not very specific • Approximately 50% of BC may express S100 • DDx for cytokeratin+/S100+ tumor inc. melanoma and BC MNF CK7 116 • Additional markers mandatory to distinguish CK18 CK7

Primary Melanoma Metastatic Melanoma

Safadi et al., Oral and Maxillofacial Pathol 2016;121:510‐519 Dwarakanath Hum Pathol 1987;18:1144‐1148 Stroup Hum Pathol 1988;19:949‐953

3 3/29/2017

Case #2 History Left Axillary Lymph Node Biopsy

• 31 yo female h/o left shoulder melanoma ; Negative left axillary sentinel nodes • Presented 7 mo later with left axillary mass • Core needle biopsy performed

Left Axillary Mass Immunohistochemistry Diagnosis CK 7 CAM5.2 GATA3 CK AE 1/3 • Metastastic carcinoma, most consistent with breast cancer

Melan‐A SOX10 S100 HMB45 • AR‐, likely ruling out adnexal carcinoma

4 3/29/2017

Melanoma Left Shoulder

Immunohistochemistry Left Shoulder Mass CK7 AE 1/3 GATA3 AR

HMB 45 S100 MelanA SOX10

BC vs Melanoma Immunohistochemistry

Case #2, Diagnosis #2 Case #1 Case #2 Melanoma BC Melanoma AE 1/3 + patchy weak‐mod + diffuse weak ‐ Left Shoulder CK7 ‐ + diffuse strong ‐ Breast ER ‐‐ND Malignant melanoma with intratumoral GATA3 ‐ + diffuse strong ‐ CK 8/18 ‐ ND ND focus of carcinoma consistent with S100 + ‐ + metastasis from breast cancer SOX10 + ‐ + Melanoma MITF + ND ND MelanA + ‐ + HMB45 + ‐ + GI CDX2 ‐ ND ND HER2 ND ‐ ND

5 3/29/2017

GATA3 Expression GATA3 in the Breast Primary Clinical Use Other Carcinomas SCC ‐ variety of sites 80% • with role in differentiation of mammary luminal Breast carcinoma (92% primary, 96% met) Adnexal carcinomas 70% epithelial cells Salivary gland carcinomas >99% Urothelial carcinoma (91% primary) Choriocarcinoma and trophoblastic tumors >99% • In BC cell lines and mouse models 1) reduces tumor initiating capacity Mesothelioma 58% of luminal progenitor cells & 2) impedes EMT by interaction with Chromophobe carcinoma 51% BRCA1 Pancreatic ductal carcinoma 10‐37% Other adenocarcinomas < 10% • In ER+ cell lines, required for estradiol stimulation of cell cycle prog lung • In ER-/BRCA1 mutant cell lines, BRCA1 unable to associate with stomach endometrium GATA3 leading to a drug-resistant and mesenchymal phenotype ovary • GATA3 somatic mutations identified most commonly in ER+ but also thyroid ER- Metastatic melanoma 0/74

Kouros‐Mehr Curr Open Cell Bio 2008;20:164 Asch‐Kendrick Hum Path 2016;48:37‐47 Miettinen Am J Surg Pathol 2014;38:13‐22 Yang Mod Pathol 2010;23:654‐61 Asch‐Kendrick Hum Path 2016;48:37‐47 Tkocz Oncogene 2012;31:3667‐78

GATA3 Expression in Breast Cancer Recommendations for GATA3 use Primary BC MBC Breast Primary Metastasis ER+ 80-100% All 73-95% • Highly sensitive for ER+, routinely • Strongly supportive of breast unnecessary origin in patient with known ER+ >99% NST (ductal) 91-100% • Greatest utility, ER‐ HG tumor, esp if history of ER+ or ER‐ BC HER2+ >99% Lobular 80-100% lacking an in situ component, positivity • Suggests mammary origin in TNBC 56% (44-70%) ER+/HER2+ 100% strongly supports breast primary, CK7+/CK20‐ carcinoma of unk further strengthened by other markers‐ primary, req other markers to ER-/HER2+ 79-88% diffuse strong CK7+, SOX10+ r/o other CK7+ adeno, TTF‐1 and • TNBC 2.6%-83%(70-80%) More sensitive and specific than napsin ‐ lung, PAX8‐renal & gyn, Useful internal positive controls ‐ mammoglobin and GCDFP‐15 for dx BC CDX2‐GI Apocrine 83-91% B9 luminal cells and lymphocytes esp in TNBC • GATA3+/CK7‐ broader ddx SCC Metaplastic 17-56% • Rules out melanoma (at present)

Asch‐Kendrick Hum Path 2016;48:37‐47 Cimino‐Mathews Hum Pathol 2013;44:1341‐9 Ciocca Hum Pathos 2009;40:489‐95 Asch‐Kendrick Hum Path 2016;48:37‐47 Miettinen Am J Surg Pathol 2014;38:13‐22 Voduc Cancer Epi Bio Prev 2008;17:365‐73 Yang Mod Pathol 2010;23:654‐61 Krings Hum Pathol 2014;45:2225‐32 Liu Am J Clin Patol 2014;141:648‐55 Miettinen Am J Surg Pathol 2014;38:13‐22

Case #3 History and Presentation Breast Core Needle Biopsy • 74‐year‐old female presented to her PCP with an inverted nipple • PMH Right elbow melanoma • Mammogram – • Right breast, 12 o' • 1.7 cm x 6.4 Photocm asymmetric Album density • 2 cm from the nipple.by Sanders, Melinda • Ultrasound 1.5 cm x 1.3 cm x 1.7 cm hypoechoic mass with posterior shadowing • A right axillary ultrasound also revealed multiple morphologically abnormal lymph nodes. • Ultrasound‐guided core needle biopsy performed.

6 3/29/2017

Hormone Immunohistochemistry HER2 FISH

Not Amplified ER (3+, 100%) PR (2+, 85%) HER2/Cep17 ratio 1.0 Average HER2 copy # 1.9

Wire-Localized Partial Mastectomy

7 3/29/2017

surgery surgery

surgery

Immunohistochemistry Breast Diagnosis ER CK7 CK8/18 GATA3 Synaptophysin • Invasive mammary carcinoma , no special type with neuroendocrine differentiation • WHO 2012 : well-differentiated [carcinoid-like] neuroendocrine carcinoma • 24 mm MelanA S100 MITF SOX10 • “Metastatic tumor” involving 11 of 24 lymph nodes, largest focus measuring 35 mm with extracapsular extension.

8 3/29/2017

Axillary Lymph node with Metastatic Breast Cancer

Axillary lymph node CK 8/18 CK 7 ER GATA3

MelanA S100 SOX10 MITF

Axillary Lymph Node Axillary Lymph node with Metastatic Melanoma

CK8/18 CK7 ER GATA3

MelanA S100 SOX10 MITF

Axillary Axillary Lymph Node with Metastatic BC and Melanoma Lymph Node GATA3 CK8/18

MelanA Sox10

9 3/29/2017

Diagnosis BC vs Melanoma Immunohistochemistry Case #1 Case #2 Case #3 • Invasive mammary carcinoma , no special type Melanoma BC Melanoma BC Melanoma AE 1/3 + patchy weak‐mod + diffuse weak ‐ + ‐ with neuroendocrine differentiation CK7 ‐ + diffuse strong ‐‐ ‐ • Metastatic mammary carcinoma involving 11 of 24 Breast ER ‐‐ND + ‐ GATA3 ‐ + diffuse strong ‐ + ‐ lymph nodes, largest focus measuring 35 mm with CK 8/18 ‐ ND ND + ‐ extracapsular extension. S100 + ‐ + ‐ + SOX10 + ‐ +Focal LN+ + • Metastatic melanoma involving 2 of 24 lymph Melanoma MITF + ND ND ‐ + nodes, 1.2 mm 0.5 mm. MelanA + ‐ +Focal LN+ + HMB45 + ‐ +ND+ GI CDX2 ‐ ND ND ND ND HER2 ND ‐ ND ‐ ND

SOX10 Expression in Breast Cancer Immunohistochemical labeling of Sox10 in breast carcinoma Intensity of Sox10 staining Total Tumor type n 0 1+2+3+4+positive • Transcription factor ‐ mediates differentiation of neural crest‐derived Luminal A212100000 (0%) cells Luminal B7600011 (14%) • Most common clinical use –IHC, support diagnosis of malignant Her2141300011 (7%) melanoma Basal‐like 32 10 0 3 4 15 22 (69%)*

• Expressed by mammary TNBC,NOS133022610 (77%)* myopepithelial cells and approximately 40% of breast cancers Metaplastic13700246 (46%)* • Most commonly expressed in TNBC Table 2 Cimino‐Mathews Hum Pathol 2013; 44:959‐965 88% (35/40) *P < .0001

Nuclear Sox10 labeling defined as 0, 1+ (1%‐25%), 2+ (25%‐50%), 3+ (50%‐75%), 4+ (>75%) Subtypes determined by surrogate panel of IHC: LumA = >75% ER+/HER2‐ Cimino-Mathews Human Pathol 2013;44:959-965 LumB = ER+/HER2+, Mollaaghababa Oncogene 2003;22:3024-3034 Basal TNBC = ER‐/PR‐/HER2‐/ CK5/6 +/or EGFR+ Dravis et al. Cell Reports 2015;12:2035–2048 TNBC, NOS = ER‐/PR‐/HER2‐/CK5‐6‐/EGFR‐ Nonaka Am J Surg Pathol 2008;32:1291-8

10 3/29/2017

Summary- Summary- What I learned from 3 cases and more than 3 antibodies: What I learned from 3 cases and more than 3 antibodies:

• Melanoma - 2nd most common non-hematopoietic neoplasm to • Approx 10% BC CK7‐, further highlighting the unreliability of CKs in this distinction metastasize to the breast, and although rare, primary melanomas of • GATA3 most sensitive marker for BC in both in breast and metastases, ER+ and ER‐, the breast occur most helpful in identification of CK7‐ TNBC + • MM must be in ddx for woman presenting with an ER-/PR- breast or • GATA3 excludes melanoma • Important pitfall, SOX 10 expressed in 92% of melanomas also expressed in 40% axillary mass, esp if unusual morph, nested and TILs breast cancers, most common in TNBC (>60%), inc. metaplastic carcinomas • Evidence of CK expression in a breast/axillary mass does not R/O MM • S100 most sensitive marker for MM (>95%), however, up to 50% of BC may express • Aberrant CK expression (esp. CK7/8/18, AE1/3, CAM5.2, MNF-116) in S100, thus ddx for S100+/SOX10+ neoplasm must inc. BC requiring additional w/u melanoma, especially metastatic, should prompt use of a battery of • When evaluating metastatic biopsy, do not underestimate value of obtaining block melanoma markers- MITF, HMB-45, MART-1 before excluding MM of primary for comparison IHC testing • Consider molecular testing in unresolvable cases

THANK YOU

11