NY-ESO-1 expression in synovial sarcoma and other mesenchymal tumors: significance for NY-ESO-1-based targeted therapy and differential diagnosis

A promising targeted therapy against NY-ESO-1 (CTAG 1B) using genetically modified T-cells in synovial sarcomas was recently demonstrated in a clinical trial at the NCI. To investigate the role of NY-ESO-1 immunohistochemistry in patient selection and gain better insight into the incidence of NY-ESO-1 expression in synovial sarcomas and other mesenchymal tumors, we evaluated NY-ESO-1 expression by immunohistochemistry in 417 tumors. This collection of samples included: 50 SS18/SSX1/2 fusion positive synovial sarcomas, 155 gastrointestinal stromal tumors (GIST), 135 other spindle cell sarcomas as well as 77 other sarcomas (chondrosarcoma, osteosarcoma, dedifferentiated liposarcoma, alveolar soft part sarcoma, rhabdomyosarcoma, angiosarcoma, malignant mesothelioma, and Ewing's sarcoma). We report that 76% of synovial sarcomas expressed NY-ESO-1 in a strong and diffuse pattern (2-3+, >50-70% of tumor cells). In contrast, only rare cases of other spindle cell mesenchymal tumor expressed NY-ESO-1 (GIST (2/155), malignant peripheral nerve sheath tumors (1/34), and dermatofibrosarcoma protuberans (2/20)). Individual cases of other sarcomas (angiosarcoma, malignant mesothelioma, chondrosarcoma, osteosarcoma, dedifferentiated liposarcoma, alveolar soft part sarcoma, and Ewing's sarcoma) were positive for NY-ESO-1. However, no positive cases were identified amongst our cohort of leiomyosarcomas (0/24), hemangiopericytoma/solitary fibrous tumors (0/40), and cellular schwannomas (0/17). In summary, we find that NY-ESO-1 is strongly and diffusely expressed in a majority of synovial sarcomas, but only rarely in other mesenchymal lesions. Beyond its role in patient selection for targeted therapy, immunohistochemistry for NY-ESO-1 may be diagnostically useful for the distinction of synovial sarcoma from other spindle cell neoplasms.     

High-grade extraskeletal myxoid chondrosarcoma: a high-grade epithelioid malignancy

AIMS: Extraskeletal myxoid chondrosarcoma is typically a low-to-intermediate grade sarcoma that is associated with a prolonged clinical course. High-grade forms are rare and not well characterized. In this series we report the clinicopathological, immunohistochemical and ultrastructural findings in four cases of high-grade extraskeletal myxoid chondrosarcoma. METHODS AND RESULTS: The patients were three men and one woman (ages 34-73 years) with tumours located in the thigh (two cases), paraspinal soft tissue and perineum. Three patients had metastases, one at 12 weeks, one at 10 months, and one at presentation of recurrent tumour. In the latter case the original tumour was low grade and became high grade when it recurred 3.5 years later. All three patients died of disease. One patient was lost to follow-up. The most striking histological feature in all four tumours was the presence of numerous large epithelioid cells. These cells were arranged in cords within myxoid matrix and in sheets devoid of matrix. Two tumours had areas of conventional extraskeletal myxoid chondrosarcoma intermixed with the high-grade areas. One tumour showed transition to high-grade spindle cell sarcoma. One tumour had cells with rhabdoid features. Immunohistochemically, two tumours focally expressed S100 protein, and one focally expressed EMA. All were negative with cytokeratin, desmin, smooth muscle actin, HMB45, CD31 and CD34. Ultrastructural features in three cases were compatible with chondrosarcoma; one tumour had aggregates of microtubules within rough endoplasmic reticulum, a characteristic feature of this tumour. CONCLUSIONS: High-grade extraskeletal myxoid chondrosaroma is a rare and aggressive soft tissue sarcoma, and should be included in the differential diagnosis of other epithelioid malignancies.     

Immunostaining for SYT protein discriminates synovial sarcoma from other soft tissue tumors: analysis of 146 cases.

Synovial sarcoma in its classic biphasic form can be distinguished readily from other soft tissue lesions; however, monophasic and poorly differentiated forms are diagnostically more problematic. For this reason, we assessed the efficacy of immunostaining for SYT and SSX1 proteins, the gene products resulting from unique synovial sarcoma translocation, to distinguish synovial sarcoma from other soft tissue lesions. A total number of 146 cases were analyzed, including 47 synovial sarcoma cases (all of which were verified by FISH to have t(X; 18) translocation and SYT-SSX fusion gene) and 99 soft tissue tumors of various types. A polyclonal IgG antibody against SYT was used to stain formalin-fixed paraffin embedded tissues. Forty-one out of 47 (87%) synovial sarcoma displayed strong positive nuclear staining (ranging from 80 to 90% of the tumor cells) for SYT antibody. Nineteen of 99 (19%) non-synovial sarcoma cases showed variable nuclear and cytoplasmic staining with SYT, which ranged from 20 to 60% of tumor nuclei, and included malignant peripheral nerve sheath tumor (5/25), solitary fibrous tumor (2/14), Ewing sarcoma (2/6), low grade fibromyxoid tumor (2/4), extraskeletal mesenchymal chondrosarcoma (2/6), gastrointestinal tumor (4/17), epithelioid sarcoma (2/2). The remaining non-synovial sarcomas were negative. This is the first study demonstrating SYT protein expression in tissue sections of synovial sarcoma. This method could provide an easy, rapid and widely applicable means of assisting in the diagnosis of synovial sarcoma, particularly when material and/or resources are unavailable for PCR or FISH-based testing. However, as variable weak staining for SYT may be encountered in a small percentage of non-synovial sarcoma sarcomas, a positive interpretation should be made only when the staining is strong, nuclear and present in the majority of cells.     

Extraskeletal myxoid chondrosarcoma: a light microscopic, immunohistochemical, ultrastructural and immuno-ultrastructural study indicating neuroendocrine differentiation

AIMS: Extraskeletal myxoid chondrosarcoma is a rare low-grade soft-tissue sarcoma with locally aggressive and metastasizing potential. Extraskeletal myxoid chondrosarcoma has distinctive clinical, light microscopic, immunophenotypic, cytogenetic and ultrastructural features. Evidence that extraskeletal myxoid chondrosarcoma often shows neuroendocrine features was first provided by Chhieng et al. on the basis of an immunohistochemical and ultrastructural study of seven cases. Our study aims to further confirm by immunohistochemistry and ultrastructural studies, including immunoelectron microscopy, that extraskeletal myxoid chondrosarcoma indeed may show neuroendocrine differentiation. METHODS AND RESULTS: Fifteen cases of extraskeletal myxoid chondrosarcoma and seven control cases of skeletal chondrosarcomas were studied. Extensive immunohistochemical analysis was performed in all cases and ultrastructural studies were done in 11 extraskeletal myxoid chondrosarcomas and three skeletal chondrosarcomas. Immunoelectron microscopy was performed on one case each of extraskeletal myxoid chondrosarcoma and skeletal chondrosarcoma. Extraskeletal myxoid chondrosarcomas expressed neuron-specific enolase (100%), synaptophysin (87%), S100 (50%), PGP 9.5 (40%), and epithelial membrane antigen (25%). Co-expression of synaptophysin and PGP 9.5 was observed in six tumours. Skeletal chondrosarcomas showed expression of S100 protein, vimentin and neuron-specific enolase in all cases. Synaptophysin, chromogranin and PGP 9.5 were not expressed in any skeletal chondrosarcoma case. Ultrastructurally, extraskeletal myxoid chondrosarcoma was characterized by distinct cords of cells immersed in a glycosaminoglycan-rich matrix. The cells were rich in mitochondria, had well-developed Golgi apparatus and there were numerous smooth vesicles. In three cases there were easily found 140-180 nm diameter membrane-bound dense-core granules in cell bodies and in processes, unrelated to the Golgi, compatible with neurosecretory granules. Fewer such granules were present in the remaining extraskeletal myxoid chondrosarcoma cases, three of which also contained intracisternal tubules typical of extraskeletal myxoid chondrosarcoma. The skeletal chondrosarcomas had scalloped cell surfaces, prominent rough endoplasmic reticulum focally distended with secretory product, and lacked neurosecretory granules. Intermediate filaments were prominent in both extraskeletal myxoid chondrosarcoma and skeletal chondrosarcomas. Immunoelectron microscopy showed synaptophysin expression in the extraskeletal myxoid chondrosarcoma but not in the skeletal chondrosarcoma case. CONCLUSIONS: This study confirms that a substantial proportion of extraskeletal myxoid chondrosarcomas show immunophenotypic and/or ultrastructural evidence of neuroendocrine differentiation, and are unlikely to be related to conventional skeletal chondrosarcomas.     

Calcium pump expression in human bone and soft tissue tumours

Ninety-one cases of human bone and soft tissue tumours were studied for calcium pump expression by strepto-avidin-biotin immunohistochemical staining with a monoclonal antibody against sarcoplasmic reticulum calcium-ATPase (mAb6F5). Two out of 5 cases of embryonal rhabdomyosarcoma, 1 out of 5 cases of biphasic synovial sarcoma, 4 of 4 cases of chordoma and all of 3 chondrosarcoma cases were positive for mAb6F5. Although this novel monoclonal antibody can be used as a marker of myogenic tumours, the present positive result for endoplasmic reticulum calcium-ATPase (calcium pump) in other tumours including chordoma, chondrosarcoma and synovial sarcoma indicates a wider immunoreactivity. The findings further suggest that intracellular calcium may play an important role in cell proliferation and/or differentiation.     

Effusion cytology in the diagnosis of malignant epithelioid and biphasic pleural mesothelioma

We reviewed the cytologic findings in pleural effusions of 36 patients in whom a diffuse epithelioid or biphasic malignant mesothelioma of the pleura developed between 1978 and 1988. Malignant neoplasms were diagnosed in 26 (72%) of the 36 patients. A specific diagnosis of a mesothelioma was rendered or suspected in 64% (23/36 patients). Mesothelioma was favored over adenocarcinoma in 81% (29/36) of patients with positive fluid cytologic findings. The contribution of effusion cytology to the final diagnosis and patient treatment was assessed. These data suggested that a cytologic diagnosis contributed useful information in most patients with malignant epithelioid and biphasic pleural mesotheliomas.     

Translocations involving chromosomes 2 and 13 in benign and malignant cartilaginous neoplasms

Cytogenetic analysis of a chrondromyxoid fibroma of bone and an extraskeletal myxoid chondrosarcoma revealed clonal chromosomal rearrangements involving chromosomes 2 and 13. The significance of these findings in these neoplasms of cartilaginous origin, one benign and one malignant, is discussed.     

Myxoid variant of adrenocortical carcinoma: report of a unique case

Myxoid variant of adrenocortical carcinomas (ACC) are rare, there being only 11 cases in the literature to date. Reported herein are the findings of a case, which in contrast to all previously reported myxoid ACC, was devoid of typical non-myxoid areas. The patient was a 61-year-old man in whom a left adrenal mass was detected during investigation of Cushing's syndrome. The adrenal was replaced by malignant cells and expanses of myxoid material. The cells were positive for melan-A, synaptophysin, vimentin and alpha-inhibin. The ultrastructural features of the cells were typical of adrenal cortical differentiation. The differential diagnosis of myxoid ACC includes extraskeletal myxoid chondrosarcoma, chordoma, myxoid adenocarcinoma, myxoma, lipomatous tumors, nerve sheath tumors, smooth muscle tumors, gastrointestinal stromal tumor and other sarcomas. The presence of myxoid material in a retroperitoneal lesion raises a broad differential diagnosis in which myxoid adrenocortical neoplasms should be included. Clinicoradiological correlation may be helpful, but special stains, immunohistochemistry and ultrastructural examination may be necessary to establish the diagnosis.     

Primary peritoneal mesotheliomas in children: a clinicopathological and immunohistochemical study of eight cases

Aims:  To present eight cases of primary diffuse peritoneal malignant mesothelioma in children <15 years old, with a discussion of the pitfalls of this diagnosis in the paediatric age group.
Methods and results:  The cases were selected based on the following criteria: (i) primary peritoneal neoplasms confined grossly or radiographically to the abdominal cavity; (ii) negative history of previous or another associated malignancy; (iii) histopathological confirmation. All patients (five female, three male) presented clinically with symptoms of abdominal pain, distention and ascites. Grossly, the tumours showed multiple, diffuse peritoneal nodules. Histologically, seven cases corresponded to epithelioid mesotheliomas and one case displayed biphasic (epithelioid and spindle) cellular proliferation. Immunohistochemical studies for cytokeratin (CK) 5/6, calretinin and low-molecular-weight CK (CAM5.2) showed strong cytoplasmic positivity in the neoplastic cells. Three patients were treated by chemotherapy. On clinical follow-up, four patients with epithelioid mesotheliomas were alive and well from 12 to 18 months after initial diagnosis; one patient with a mixed (biphasic epithelioid/sarcomatoid) mesothelioma died of tumour 24 months after diagnosis.
Conclusions:  Peritoneal malignant mesothelioma in children is a rare condition that can introduce difficulties in histopathological diagnosis.     

Monophasic synovial sarcoma, epithelioid sarcoma and chordoid sarcoma: ultrastructural evidence for a common histogenesis, despite light microscopic diversity

Ultrastructural examination of six rare sarcomas-four monophasic spindle cell tumours, one epithelioid sarcoma, one chordoid sarcoma-has revealed marked similarities at the electron microscopic level despite widely divergent light microscopic appearances. These features consisted of: 1 the presence of two cell types, viz. a clear cell and a cell resembling the fibroblast; 2 pseudoglandular spaces with projecting microvilli or filopodia, and with related tight junctions; 3 an amorphous intercellular ground substance with focal condensation into recognizable basement membrane. The findings suggest a common maturation of these diverse tumours to synovial-like tissue, and support the proposal of Hajdu Shiu & Fortner (1977) that these be considered variants of synovial sarcoma. Published ultrastructural studies of synovial, epithelioid and chordoid sarcoma are reviewed in the light of these findings. The ultrastructural differentiation of synovial sarcoma from extra-skeletal myxoid chondrosarcoma, chordoma and the spectrum of malignant spindle cell tumours is discussed.     

HLA-DR (Ia-like) reactivity in tumors of bone and soft tissue: an immunohistochemical comparison of monoclonal antibodies LN3 and LK8D3 in routinely processed specimens.

Recent studies of Class II histocompatibility antigen expression in bone and soft tissue sarcomas have suggested that malignant fibrous histiocytoma (MFH) may express HLA-DR, whereas histologically similar pleomorphic, epithelioid, and spindle cell malignant neoplasms generally do not. To test whether these observations are reproducible in the differential diagnosis of soft tissue sarcomas, anti-HLA-DR antibodies LK8D3 and LN3 were applied to formalin-fixed, paraffin-embedded sections of MFH, neurofibrosarcoma (NFS), leiomyosarcoma (LMS), synovial sarcoma (SS), fibrosarcoma (FS), angiosarcoma (AS), Kaposi's sarcoma (KS), chondrosarcoma (ChS), "dedifferentiated" chondrosarcoma (DChS), osteosarcoma (OS), epithelioid sarcoma (ES), and clear cell sarcoma (CCS; malignant melanoma of soft parts). The only consistent difference in Class II antigen expression was seen in the group of neoplasms composed of large polygonal cells. Among the latter lesions, four of six clear cell sarcomas were labeled by LK8D3 or LN3, but none of 12 epithelioid sarcomas were reactive. Otherwise, a diversity of tumors in other morphologic categories expressed Class II antigens, with no clear diagnostic patterns. These results may be of use in the diagnostic separation of large cell epithelioid tumors of soft tissue, but neither LN3 nor LK8D3 appears to be helpful in the identification of other sarcomas.     

Extra-axial chordoma

Chordomas are low-grade malignant tumors of bone that occur almost exclusively in the axial skeleton. Other neoplasms with a similar histologic picture but an extra-axial location have been described, including parachordoma, myxoid chondrosarcoma, and extra-axial chordoma. We herein present another case of the rare extra-axial chordoma. A 41-year-old woman developed an 8.3 cm mass in the pubic bone. The gross, microscopic, and immunohistochemical findings were identical to those of a classic chordoma. Parachordoma and myxoid chondrosarcoma were excluded from the differential diagnosis. Five previously reported cases of extra-axial chordoma were reviewed and found also to demonstrate clinical and pathologic features specific to chordoma, despite arising in an extra-axial location. Although rare, extra-axial chordoma does exist and should be recognized and managed in a similar fashion to its well-described counterpart. It must be differentiated from other histologic mimics, because the treatment and prognosis can differ significantly.     

Biphasic solitary fibrous tumour: a report of two cases with epithelioid features

We present two cases of solitary fibrous tumour (SFT) showing biphasic morphology with a spectrum of malignant epithelioid components. Slides prepared from formalin-fixed and paraffin-embedded tissue from both cases were stained with haematoxylin and eosin and by immunohistochemistry. Interphase fluorescent in situ hybridisation studies were performed in both cases using paraffin-embedded tissue to look for the t(X;18) translocation, thereby to exclude synovial sarcoma. Both cases showed biphasic morphology with some areas having typical benign spindled SFT morphology (including CD34 expression) and other areas having a malignant epithelioid appearance. In one of the cases, the epithelioid area, which was well circumscribed and showed packeting of cell groups, demonstrated expression of cytokeratin and epithelial cadherin but not of CD34. In the second case, the immunophenotype of the epithelioid component was similar to that of the benign SFT component. These findings suggest that epithelioid change in SFT shows a range of differentiation at one end, similar to that of a standard SFT, and at the other end, possibly acquiring epithelial characteristics.     

TLE1 expression in malignant mesothelioma

Malignant mesothelioma, an aggressive and often lethal tumor commonly associated with asbestos exposure, has been morphologically classified into epithelial, biphasic, and sarcomatoid subtypes. Histological distinction between biphasic or sarcomatoid mesothelioma and synovial sarcoma may be problematic in certain circumstances of intrathoracic location because of their similar clinicopathologic features, including not only their morphology but also occasional positive immunoreaction of mesothelioma markers. TLE1, which plays an important role in Wnt pathway, has been shown to be a specific marker for synovial sarcoma and diagnostically is useful; however, TLE1 expression in malignant mesotheliomas has not been fully evaluated. We immunohistochemically examined the expression of TLE1, factors related to the Wnt pathway including β-catenin and cyclin D1, and mesothelioma markers including calretinin, HBME-1, cytokeratin 5/6, and thrombomodulin in 29 malignant mesotheliomas. TLE1 was variably expressed in 28 malignant mesotheliomas regardless of histomorphological subtype with >25% of positive cells in 20 cases (69.0%). There was no evidence of association of TLE1 expression with immunoreactivity to other markers. Our study showed no or limited value of the immunohistochemical TLE1 expression in distinguishing malignant mesothelioma and synovial sarcoma.     

p63 immunohistochemical staining is limited in soft tissue tumors

p63 is a p53 homolog that is expressed in various normal epithelial tissues and epithelial malignancies. Its expression in mesenchymal lesions has not been examined in depth; therefore, we studied p63 expression by immunohistochemical analysis in 650 soft tissue tumors. We found that p63 expression is limited in soft tissue tumors. The majority of tumors studied were p63-, including all cases of angiosarcoma, lipomatous neoplasms, dermatofibrosarcoma protuberans, solitary fibrous tumor, schwannoma, neurofibroma, gastrointestinal stromal tumor, and leiomyosarcoma. Nuclear p63 reactivity was found in a subset of soft tissue myoepithelioma and myoepithelial carcinoma of soft tissue, cellular neurothekeoma, soft tissue perineurioma, Ewing sarcoma/peripheral neuroectodermal tumor, diffuse-type giant cell tumor, and giant cell tumor of soft parts. Infrequent, weak, or focal p63-staining patterns were observed in low-grade fibromyxoid sarcoma, malignant peripheral nerve sheath tumor, extraskeletal myxoid chondrosarcoma, myxofibrosarcoma, proximal-type epithelioid sarcoma, synovial sarcoma, embryonal rhabdomyosarcoma, desmoplastic small round cell tumor, atypical fibroxanthoma, and spindle cell melanoma. Absent p63 expression is typical for most soft tissue tumors, including most (but not all) that would be in the differential diagnosis of spindle cell squamous carcinoma.     

Absence of expression of SMARCB1/INI1 in malignant rhabdoid tumors of the central nervous system, kidneys and soft tissue: an immunohistochemical study with implications for diagnosis.

Malignant rhabdoid tumors are high-grade neoplasms of the central nervous system (CNS), kidneys and soft tissue that usually occur in children. The histologic diagnosis of malignant rhabdoid tumor depends on identification of characteristic rhabdoid cells-large cells with eccentrically located nuclei and abundant, eosinophilic cytoplasm-and immunohistochemistry with antibodies to vimentin, keratin and epithelial membrane antigen. In most malignant rhabdoid tumors, the SMARCB1/INI1 gene, located in chromosome band 22q11.2, is inactivated by deletions and/or mutations, so genetic diagnosis is often possible. However, tissue may not be available for genetic analysis or studies not confirmatory. We assessed SMARCB1/INI1 expression in 17 rhabdoid tumors and 57 other tumors of the CNS, kidney or soft tissue using immunohistochemistry. In total, 12 brain, three renal and two soft tissue rhabdoid tumors were examined along with four glioblastomas, four pilocytic astrocytomas, four oligodendrogliomas, two ependymomas, two choroid plexus papillomas, five pituitary adenomas, four germinomas, four renal carcinomas with Xp11.2 translocations, two clear cell sarcomas, two Wilms' tumors, one renal medullary carcinoma, two desmoplastic small round cell tumors, two alveolar rhabdomyosarcomas, two embryonal rhabdomyosarcomas, one low-grade chondrosarcoma, two extraskeletal myxoid chondrosarcomas, one mesenchymal chondrosarcoma, four malignant peripheral nerve sheath tumors, five metastatic carcinomas and four epithelioid sarcomas, two primary and two metastatic. The neoplastic cells of all rhabdoid tumors, the four epithelioid sarcomas and the renal medullary carcinoma did not express SMARCB1/INI1 by immunohistochemistry; neoplastic cells of all other tumors expressed SMARCB1/INI1. Immunohistochemistry to assess expression of SMARCB1/INI1 may be useful in the diagnosis of rhabdoid tumors of the CNS, kidneys and soft tissue.     

Chondroides Lipom Klinisch-pathologische, immunhistochemische und ultrastrukturelle Analyse von 6 Fällen einer distinkten Entität im Spektrum der Lipome

A series of six cases of chondroid lipoma is reported in this paper. The age of the patients, four women and two men, ranged from 34 to 75 years. Four tumours arose in the lower and two in the upper extremities between 4.5 and 8 cm in size. Whereas two neoplasms were located in the subcutis, four lesions were seen in perifascial or intramuscular tissues. Histologically, all neoplasms were encapsulated and characterized by a somewhat lobular growth pattern. The neoplasms were composed of mature adipocytes, uni - and multivacuolated lipoblasts, and nests and strands of cells reminiscent to hibernoma cells or chondrocytes. Immunohistochemically and ultrastructurally the lipogenic nature of the latter type of cells was confirmed. Interestingly, focal immunopositivity of tumour cells for cytokeratin was found in two cases. Tumour cells were set in a myxohyaline matrix showing different degrees of degeneration. Chondroid lipoma represents a distinct entity in the spectrum of lipomatous lesions and has to be distinguished from benign and malignant neoplasms (extraskeletal chondroma, hibernoma, lipoblastoma, chondrolipoangioma, myoepithelioma, myxoid/round cell liposarcoma, and extraskeletal myxoid chondrosarcoma). Because they are easily misdiagnosed as malignant tumours, familiarity with the features of chondroid lipoma is of practical importance to avoid an overtreatment.     

Usefulness of p16/CDKN2A fluorescence in situ hybridization and BAP1 immunohistochemistry for the diagnosis of biphasic mesothelioma

Malignant mesothelioma is a highly aggressive neoplasm, and the histologic subtype is one of the most reliable prognostic factors. Some biphasic mesotheliomas are difficult to distinguish from epithelioid mesotheliomas with atypical fibrous stroma. The aim of this study was to analyze p16/CDKN2A deletions in mesotheliomas by fluorescence in situ hybridization (FISH) and BAP1 immunohistochemistry to evaluate their potential role in the diagnosis of biphasic mesothelioma. We collected 38 cases of pleural mesotheliomas. The results of this study clearly distinguished 29 cases of biphasic mesothelioma from 9 cases of epithelioid mesothelioma. The proportion of biphasic mesotheliomas with homozygous deletions of p16/CDKN2A in total was 96.6% (28/29). Homozygous deletion of p16/CDKN2A was observed in 18 (94.7%) of 19 biphasic mesotheliomas with 100% concordance of the p16/CDKN2A deletion status between the epithelioid and sarcomatoid components in each case. Homozygous deletion of the p16/CDKN2A was observed in 7 (77.8%) of 9 epithelioid mesotheliomas but not in fibrous stroma. BAP1 loss was observed in 5 (38.5%) of 13 biphasic mesotheliomas and in both epithelioid and sarcomatoid components. BAP1 loss was observed in 5 (62.5%) of 8 epithelioid mesotheliomas but not in fibrous stroma. Homozygous deletion of p16/CDKN2A is common in biphasic mesotheliomas, and the analysis of only one component of mesothelioma is sufficient to show that the tumor is malignant. However, compared with histology alone, FISH analysis of the p16/CDKN2A status and BAP1 immunohistochemistry in the spindled mesothelium provide a more objective means to differentiate between biphasic mesothelioma and epithelioid mesothelioma with atypical stromal cells.