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A 47 year old female with multiple lung nodules Disclosure of Relevant Financial Relationships Tamar Giorgadze, MD, PhD Professor of Pathology Medical College of Wisconsin Milwaukee, Wisconsin USCAP requires that all planners (Education Committee) in a position to influence or control the content of CME disclose any relevant financial relationship WITH COMMERCIAL INTERESTS which they or their 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. Index case: Clinical History I have nothing to disclose A 47 year old female (never smoker) presented with abdominal pain CT scan revealed multiple bilateral lung nodules CT guided fine needle aspiration (FNA) and core biopsy of the largest 1.0 cm solid lung nodule were performed at the Outside Institution #1 The patient was referred to Weill Cornell Medicine for further evaluation The pathology slides have been reviewed within the Weill Cornell Medical College (WCMC) Department of Pathology and Laboratory Medicine FNA, direct smears, low power view FNA, direct smears DQ, 10x PAP, 10x DQ, 40x 1

FNA, high power view ThinPrep, high power view Index Case: Core Biopsy Index case: IHC Core biopsy, H&E, 40x Core biopsy, H&E, 40x CK AE1/AE3 CK 7 Index case: IHC Index case: IHC TTF 1 CD56 Synaptophysin Chromogranin 2

Index case: IHC Outside Institution #1 Diagnosis Lung, RLL, FNA and Core Biopsy: Atypical cells present, consistent with carcinoid. CD117(CKIT) CD45(LCA) Ki 67 Index case: Differential Diagnosis NET: Cytology The differential diagnosis of spindle cell lesions in the lung is broad It may include both primary and metastatic tumors with spindle cell features Multiple bilateral pulmonary nodules are due to metastatic disease in about 70% of cases In the absence of previous history of malignancy, the main DD in our case was with a neuroendocrine tumors (NET), including typical carcinoid (TC) and atypical carcinoid (AC) In the setting of multiple bilateral pulmonary nodules, consideration should have been given to diffuse idiopathic neuroendocrine cell hyperplasia Organoid growth pattern of tumor cells: Nested Rosette like Islands Ribbons Festoons Trabeculae PAP, 4x Cell block, H&E, 10x Carcinoid Tumor: Cytology NET: Cytology and IHC DQ, 10x DQ, 10x DQ, 40x NET typically shows strong immunoreactivity to one or more neuroendocrine markers Strong positivity to cytokeratins is seen in more than 80% of cases. Staining for TTF 1 in ~40% of cases TC: <2 mitoses per 2mm 2 ; <2% by Ki 67 IHC Separation of TC from AC may be difficult in cytology and small biopsy specimens Synaptophysin CD56 Cell block, H&E, 40x Chromogranin AE1/AE3 Ki 67/CD45 3

NET: Cytology and IHC Index Case : Additional IHC on de stained slides performed at WCMC AC may show nuclear molding, more frequent mitoses and/or pinpoint foci of necrosis AC: 2 10 mitoses per 2 mm 2 ; up to 20% by Ki 67 DQ, 60x AE1/AE3 DQ, 60x Cell Block, H&E, 40x DQ, 60x Ki 67/CD45 Negative: PAX 8 Synaptophysin CD10 Index Case: Summary of IHC reactivity Index Case: Review Diagnosis at WCMC Positive CD56 CD10 Ki67 (<2%) of the lesional cells Negative AE1/AE3 CK7 CK20 CAM5.2 Chromogranin Synaptophysin LCA CKIT PAX 8 Lung, right lower lobe, 1.0 cm nodule, FNA and core needle biopsy: Spindle cell lesion. Comment: Overall, the immunoprofile of the lesional cells makes the diagnosis of a neuroendocrine tumor unlikely. A precise diagnosis cannot be rendered on this limited tissue material. Recommend additional tissue sampling for further characterization of this lesion. Repeat CT and FNA at WCMC after 5 months Repeat FNA of a RLL Nodule at WCMC: IHC 10 nodules in each lung, some increased in size and some decreased The nodules that increased in size were more cystic with no definite increase in solid component The previously aspirated largest right lung nodule increased in size to 1.4 cm and now was mostly cystic Repeat FNA of this nodule was performed Cell block, H&E, 40x CD56 CD10 Chromogranin Desmin MyoD1 ER PR 4

Repeat FNA of a RLL Nodule at WCMC: IHC Positive CD56 CD10 Desmin MyoD1 (focal) ER (>50% of cells) PR (rare cells) WT1 showed cytoplasmic uptake Ki 67 demonstrated low proliferation (2 3%) Negative AE1/AE3 CAM5.2 CK7 Synaptophysin Chromogranin TTF 1 SMA CD99 S100 protein CD34 CD31 PAX 8 Inhibin Other Spindle Cell Tumors in the Lung Solitary fibrous tumor (SFT) Synovial sarcoma Peripheral nerve sheath tumor Smooth muscle tumor Sarcomatoid mesothelioma Melanoma Gastrointestinal stromal tumor (GIST) Ovarian stromal tumor Endometrial stromal tumor Solitary Fibrous Tumor: Cytology Solitary Fibrous Tumor: IHC SFT is typically immunoreactive for CD34, Bcl 2, and CD99 SFT is negative for epithelial markers DQ, 10x DQ, 40x Cell block, H&E, 10x Bcl 2 CD34 Solitary Fibrous Tumor: IHC Synovial Sarcoma: Cytology Almost all SFT harbor an NAB2 STAT6 fusion gene Nuclear expression of STAT 6 has been recently reported as a highly sensitive and specific surrogate IHC marker for SFT Cell block, H&E, 10x DQ, 40x DQ, 40x CD34 STAT 6 PAP, 10x PAP, 20x 5

Synovial Sarcoma: IHC Synovial Sarcoma: IHC IHC to support the diagnosis of synovial sarcoma (SS) include cytokeratins, CD99, Bcl 2, EMA, and CD56 The molecular test for t(x;18)(p11.2;q11.2) chromosomal translocation can help to confirm the diagnosis Positive TLE 1 (Transducin like enhancer of split 1) nuclear staining is a useful immunomarker, has a strong correlation with the t(x;18) chromosomal translocation, and can been seen in 96% 97% of cases The gold standards for this diagnosis of SS: morphology, ancillary IHC for traditional markers, and molecular confirmation of SS associated fusion genes Cell block, H&E, 20x CD99 Bcl 2 TLE 1 H&E, 20x CD99 Bcl 2 EMA Peripheral Nerve Sheath Tumors: Cytology and IHC Schwannoma Schwannomas are rarely seen in lung FNA specimens Cytologically, large fascicular or syncytial tissue fragments of spindle cells, rare singly scattered spindle cells The individual tumor cells generally show bipolar cytoplasmic processes, bent, fishhook or wavy nuclear morphology, with uniform chromatin and occasional intranuclear pseudoinclusions Schwannomas are diffusely and strongly immunoreactive for S 100 protein Malignant peripheral nerve sheath tumors (MPNST) will show more nuclear pleomorphism MPNST will show only focal weak immunoreactivity for S 100 protein, but can show positivity for CD34, GFAP, and focal staining for EMA PAP, 4x DQ, 4x DQ, 40x DQ, 40x MPNST Smooth Muscle Tumors: Cytology and IHC PAP, 10x Core biopsy, H&E, 20x S 100 Melanoma cocktail Cytology smears of benign metastasizing leiomyoma (LM), metastatic leiomyosarcoma (LMS), and lymphangioleiomyomatosis (LAM) may demonstrate singly scattered and cohesive tissue fragments of smooth muscle cells / smooth muscle like cells Smears obtained from LM are usually hypocellullar or moderately cellular with no mitotic figures or necrosis observed Low grade LMS may morphologically resemble LM The cell clusters may demonstrate a fascicular growth pattern with irregular borders and branches of delicate capillaries The cytoplasm is often scant Tumor cells with abundant eosinophilic cytoplasm, elongated bipolar wispy cytoplasmic processes, and multinucleated tumor can also be seen High grade tumors show more nuclear pleomorphism, mitotic figures and areas of necrosis 6

Leiomyosarcoma: Cytology Leiomyosarcoma: IHC DQ, 10x PAP, 10x MSA Caldesmon Desmin CD10 ER PR Sarcomatoid Mesothelioma Case courtesy: Dr. Jason Chang LAM cell clusters: (A) ThinPrep (B) Cell Block Arrow: outer flattened cells Touch Prep, DQ, 40x Core biopsy, H&E, 20x H&E, 4x H&E, 40x (A) The inner cells with strong positivity for SMA; (B) The inner cells with strong positivity for HMB 45; (C) The superficial cells showing immunoreactivity for D2 40 DQ, 60x DQ, 60x DQ, 60x CK5/6 HBME 1 Sarcomatoid Mesothelioma: IHC Melanoma Melanoma cocktail S 100 SOX10 DQ, 10x Cell block, H&E, 20x DQ, 40x H&E, 4x H&E, 40x 7

When melanoma is negative for S100: diagnostic pitfalls GIST: Cytology & IHC Riddle ND & Bui MM Arch Pathol Lab Med; Vol 136, March 2012, 238 9 AE1/AE3 Chromogranin CKIT S 100 Cytokeratin positivity has been reported in 10% to 26% of metastatic melanomas Primary and metastatic melanomas showing small cell morphology and aberrant expression of neuroendocrine markers have been reported. The small cell population is usually devoid of pigment and either completely negative or only focally positive for S100 Melanoma demonstrates immunophenotypic heterogeneity DQ, 10x PAP, 20x Cell block, H&E, 20x DOG1 SMA Granulosa Cell Tumor Cell block, H&E, 10x DQ, 10x Inhibin DQ, 40x Calretinin CD10/CD56 Coexpression: Immunoquery Discrete Diagnosis CD10 / # of cases CD56 / # of cases Neuroendocrine Tumor 25% / 20 91% / 47 Solitary Fibrous Tumor 64% / 14 0% / 1 Synovial Sarcoma 6% / 31 51% / 68 MPNST 73% / 11 23% / 26 Leiomyosarcoma 50% / 34 76% / 34 Melanoma, Spindle Cell Type 40% / 30 7% / 130 GIST, Spindle Cell Type 19% / 52 19% / 168 Granulosa Cell Tumor, Ovary 6% / 16 100% / 40 Stromal Tumor, Endometrium 81% / 42 67% / 6 Repeat FNA of a 1.4 cm RLL nodule: Diagnosis Lung, right lower lobe, 1.4 cm nodule, fine needle aspiration: Malignant spindle cell neoplasm with heterologous differentiation. Comment: Tumors of this type can be seen in the gynecologic tract and a possibility would be uterine origin. 8

Lung, RLL and RML, wedge resection at WCMC Lung, RLL, wedge resection: IHC Desmin ER PR Negative: H&E, 2x H&E, 10x H&E, 40x AE1/AE3 Ki 67 PAX 8, Myo D1, WT 1, CD34, CD31, Inhibin Lung, RLL and RML, wedge resection: Metastatic low grade endometrial stromal sarcoma with focal smooth muscle differentiation. Comment: As per additional information, the patient had a recent hysterectomy at another institution (Outside Institution #2). It would be helpful to review the corresponding pathology slides. Outside Institution #2 Diagnosis Morcellated uterus: Leiomyomata Hysterectomy Specimen from the Outside Institution #2: Pathology Review at WCMC Hysterectomy: Pathology Review Diagnosis at WCMC Morcellated uterus: One slide demonstrates myxoid and paucicellular low grade spindle cell neoplasm amidst the myometrium, similar to that seen in the lung metastasis Leiomyomata H&E, 10x H&E, 20x 9

Low grade Endometrial Stromal Sarcoma (LGESS) LGESS accounts for 0.2% of female genital tract malignancies The age range of the patients is between 40 to 58 years old Rare cases have been reported in younger women or young adolescents Patients typically present with abnormal uterine bleeding, pelvic pain and dysmenorrhea, however about 25% of patients can be asymptomatic About 30 50% of patients have extrauterine spread at the time of diagnosis LGESS rarely presents initially at an extrauterine site including metastatic disease involving the ovary and lung, causing diagnostic challenges LGESS have been rarely reported as incidental findings in uterine morcellation specimens Three patients had resected uterine endometrial stromal sarcoma (ESS) originally diagnosed as smooth muscle tumor, in one patient as uterine neoplasm resembling sex cord tumor, and in one patient ESS was recognized only after review of previous hysterectomy slides ESS should be included in the differential diagnosis of nonepithelial pulmonary neoplasms Kim GY, Sung CO, Han J, Park OJ, Lee KS Pulmonary Metastases of Uterine Endometrial Stromal Sarcoma: Diffuse Micronodular and Ground Glass Opacities: A Case Report. J Korean Med Sci 2004; 19: 901 3 The most common growth pattern of pulmonary metastatic ESS is well circumscribed nodules with entrapped air spaces lined by non neoplastic respiratory epithelium. The unusual growth patterns are solitary nodule, satellite with infiltrative margins, lymphangitic pattern and bilateral spontaneous pneumothoraces associated with predominantly cystic lesions mimicking LAM, which can contribute to the diagnostic dilemma. This is especially true if clinicians and/or pathologists are unaware of a prior diagnosis of uterine ESS. Prior misdiagnosis of a uterine tumor can also be misleading. LGESS: Ancillary Studies A useful initial IHC panel for LGESS work up is reported to be CD10, ER/PR and at least two smooth muscle In rare cases, CD10 can be negative in LGESS, and conversely smooth muscle tumors may be immunoreactive to CD10 Foci of smooth muscle or sex cord differentiation in LGESS can be immunoreactive for smooth muscle markers, inhibin, calretinin, WT1, Melan A, and CD99 It is important to consider the relative intensity and distribution of these immunostains and interpret them in view of morphologic and clinical findings The majority of LGESS harbor t(7;17)(p15;q21) translocation, which results in JAZF1 SUZ12 gene fusion Molecular testing for diagnosis of LGESS is not routinely performed It may be helpful in cases with unusual clinical presentation or morphologic findings Index Case: Summary LGESS presented initially as pulmonary nodules Outside Institution #1: Lung FNA and core biopsy have been initially misinterpreted as NET WCMC diagnosis (following review, repeat FNA, and wedge resection): Metastatic LGESS Outside Institution #2: Primary uterine LGESS has been unrecognized in the morcellated hysterectomy specimen without considering lung nodule evaluation results Thus, the patient has been rendered three different diagnosis in three different institutions 10

WHAT CAN WE LEARN FROM THIS CASE? THE IMPORTANCE OF BEEING VIGILANT Potential pitfalls in the diagnosis of metastatic LGESS in a patient without clinical history of a gynecologic neoplasm Judicious use of ancillary studies and remembering their limitations, especially while evaluating small tissue samples Significance of compiling all clinical data and correlating various surgical pathology specimens evaluation results from different institutions This case also demonstrates value of clinical care continuity in avoiding diagnostic pitfalls The vigilant eye. Giorgadze TA. Int J Surg Pathol. 2013 Feb;21(1):46 THANK YOU 11