Thymic epithelial tumors (TETs), including thymomas,

Transcription

1 Evolution of Classification of Thymic Epithelial Tumors in the Era of Dr Thomas V. Colby Anja C. Roden, MD Context. Numerous histomorphologic and staging classifications of thymic epithelial tumors (TETs) have been proposed during the last century, suggesting that the classification of these tumors is challenging and controversial. Difficulties of classifying TETs include various combinations of epithelial cells and lymphocytes and the paucity of these tumors. The prognostic significance, specifically of the histomorphologic classifications, has been debated. Early classifications were also challenged by the uncertainty of the neoplastic component(s) of the tumor. Objective. To discuss the evolution of the histomorphologic classification and the staging system of TET. Controversies and problems of some classifications and their importance for therapeutic management and prognosis will be reviewed. Classifications that incorporated new concepts and approaches at the time or outcome studies will be highlighted. Current classifications will be discussed and the staging system that was recently proposed for the upcoming eighth American Joint Committee on Cancer staging will be described. Data Sources. Search of literature database (PubMed) and current (2015) World Health Organization classification. Conclusions. Histomorphologic and staging classifications of TET have evolved during the last century and especially during the era of Thomas V. Colby, MD. Evidence supports that the staging system has prognostic implications independent of and superior to the histomorphologic classification. Histomorphology appears to be important for biologic features of TET. (Arch Pathol Lab Med. 2017;141: ; doi: / arpa ra) Thymic epithelial tumors (TETs), including thymomas, thymic carcinomas, and thymic neuroendocrine tumors, are neoplasms that in general occur in the prevascular (anterior) mediastinum. Although TETs represent the most common primary prevascular mediastinal neoplasms in adults, encompassing approximately 20% of all primary tumors in that area, they are overall infrequent. The incidence for thymomas ranges between 1.3 and 2.5 per million per year. 1 3 Thymic carcinomas are even less frequent. Thymic epithelial tumors have been reported at any age but occur most commonly at 40 to 60 years of age, and are extremely rare in children. There is no sex predilection in TETs. 1,4 6 Thymic epithelial tumors can be classified according to their histopathologic features or staged based on the presence and extent of invasion, implants, lymph node involvement, and/or distant metastases. At least 24 histomorphologic classifications and 14 staging systems have been proposed within the last century (Table 1). Accepted for publication June 2, From the Department of Laboratory Medicine and Pathology, Mayo Clinic Rochester, Rochester, Minnesota. The author has no relevant financial interest in the products or companies described in this article. Portions based on a presentation given at the 2016 Mayo Clinic Pathology Update: A Tribute to the Career of Thomas V. Colby, MD meeting; February 4, 2016; Phoenix, Arizona. Reprints: Anja C. Roden, MD, Department of Laboratory Medicine and Pathology, Mayo Clinic Rochester, 200 First St SW, Rochester, MN ( roden.anja@mayo.edu). CHALLENGES OF THE CLASSIFICATION OF THYMIC EPITHELIAL TUMORS The classification of TET has been challenging because of (1) the presence of various combinations of epithelial cells and lymphocytes that compose these tumors; (2) the paucity of these neoplasms, making large studies difficult; (3) an uncertainty about the neoplastic component(s) of TET (thymic epithelial cells and/or lymphocytes) in early classifications; (4) controversies about the prognostic significance of classifications and the malignant potential of thymomas; and (5) the diversity of thymic-derived tumors (ie, TETs, lymphomas, germ cell tumors). Many pathologists have struggled with the classification of TET. For instance, Ewing 7 noted in 1916 that no group of tumors has more successfully resisted attempts at interpretation and classification than those of the thymus. Lowenhaupt 8 stated in 1948 that thymic tumors, because of their rarity and diversity, have long resisted attempts to reach valid conclusions from [the] variously described and variously interpreted material available from the literature. In 1961, Bernatz et al 9 began their article on classification of thymoma with the remarks that a study of thymic tumors must enjoy a prominent but unenviable position in any list of frustrating experiences inherent in the pursuit of medical knowledge and that remarkable as is this dissidence all reports overwhelmingly agree that there is much confusion as well as controversy about this group of tumors. In the early years of histomorphologic classification of TETs, it was not clear whether the epithelial cells and/or the lymphocytes were the neoplastic component. In 1917, Bell Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

2 Table 1. Histomorphologic Classifications and Staging Systems of Thymic Epithelial Tumors Proposed During the Last Century Histomorphologic Classifications Staging Systems Year Author Country Year Author Country 1928 James Ewing United States 1961 Philipp E. Bernatz United States 1932 Douglas Symmers United States 1978 N. P. Bergh Sweden 1940 James Ewing United States 1978 Gerald D. Levine and Juan Rosai United States 1941 C. Alexander Hellwig United States 1979 Earle W. Wilkins and Benjamin United States 1948 Elizabeth Lowenhaupt United States Castleman 1950 William D. Seybold United States 1981 Akira Masaoka Japan 1955 Benjamin Castleman United States 1982 Groupe d Etude des Tumeurs France 1956 Lalla Iverson United States Thymiques 1956 Donald B. Effler United States 1985 Jeanne M. Verle France 1957 Raffaele Lattes United States 1991 Yosuke Yamakawa and Akira Japan 1957 A. D. Thomson United Kingdom Masaoka (TNM) 1959 George D. Andritsakis United States 1994 Kenji Koga Japan 1961 Philipp E. Bernatz United States 1994 Ryosuke Tsuchiya and Kenji Koga Japan 1965 Merle A. Legg United States (TNM) 1966 Hiroshi Watanabe Japan 1997 Saul Suster and Cesar A. Moran United States 1976 Juan Rosai and Gerald D. Levine United States 2003 Tumor-Node-Metastasis 1978 Gerald D. Levine and Juan Rosai United States 2012 Cesar A. Moran United States Mirella Marino and Hans Konrad Müller-Hermelink Thomas Kirchner and Hans Germany Germany 2014 International Association for Study of Lung Cancer/International Thymic Malignancy Interest Konrad Müller-Hermelink Group 1999 Saul Suster and Cesar A. Moran United States 1999 World Health Organization 2000 T.-T. Kuo Taiwan 2004 World Health Organization 2015 World Health Organization noted, in a case report of a thymoma, that the tumor was likely of epithelial origin. Ewing 11 mentioned in 1928 that the chief source of [lymphosarcoma or thymoma] is probably the reticulum cell, but lymphocytes are often present in abundance. Although it was still debated, in 1948, Lowenhaupt 8 remarked on the epithelial derivation of this entire group of neoplasms, seeming to conclude that the epithelial cells were the neoplastic cells. In 1972, Zeiller and Dolan 12 showed in experiments in mice and rats that thymocytes are not derived from the thymus but rather originate from the bone marrow and migrate to the thymus. These findings suggested that although they are present in close vicinity in the same organ, the embryologic origin of thymic epithelial cells differs from that of thymocytes. Rosai and Levine 13 concluded in 1976 that a simultaneous neoplastic transformation of 2 cellular components, epithelial and lymphocytic, of different embryogenesis within the same organ would be unlikely. Rather, they suggested that the epithelial cells were the neoplastic cells of thymomas because (1) the epithelial cells have morphologic and biologic properties of neoplastic cells, (2) the epithelial cells are present in the invasive tumor and metastases, (3) some thymomas are only composed of epithelial cells, and (4) in cytologically malignant thymomas the anaplastic features are restricted to [the epithelial cells]. The authors considered the lymphocytes of thymomas to be benign because (1) it had been shown that thymic lymphocytes are derived from the bone marrow, (2) light microscopic and ultrastructural features of lymphocytes in thymoma are not different from those of normal resting or stimulated lymphocytes, and (3) the lymphocytes lack biologic properties of neoplastic cells. Today the neoplastic nature of the epithelial cells and the benign/reactive properties of lymphocytes in TET have been well accepted. EVOLUTION OF THE HISTOMORPHOLOGIC CLASSIFICATION OF THYMIC EPITHELIAL TUMORS At least 24 histomorphologic classifications of TET have been proposed during the last century (Table 1). Eightyeight years ago, in 1928, Ewing 11 categorized thymic tumors into 2 main groups, the lymphosarcoma or thymoma and the carcinoma (Table 2). A third category, spindle-cell or myxosarcoma, comprised rare tumors that were thought to derive from the stroma. Although Ewing still questioned the cell of origin of the lymphosarcoma or thymoma, he concluded from his own case studies that the reticulum [epithelial] cells are the main or sole source of the tumor and the lymphocytes are largely passive but are often present in abundance. In 1948, Lowenhaupt 8 classified thymic tumors according to the stage of embryonic development of the normal thymus (Table 2). Although this classification was not used in subsequent classifications, Lowenhaupt 8 introduced some fundamental thoughts that were later confirmed in other studies, such as that TETs are of epithelial origin and that early surgical excision, if possible, is the treatment of choice. In 1955, Castleman 14 defined thymomas as tumors composed of lymphocytes and epithelial cells. Castleman did not divide thymomas into subtypes because he argued that there are so many variations within a given tumor that there are no foolproof criteria differentiating the thymoma in a patient with myasthenia gravis from one without myasthenia gravis, and that there are no microscopic differences between the thymoma that is limited to the thymic gland and the one that extends beyond its confines.... He was not entirely convinced that thymomas are malignant, even though he described that in a quarter of thymomas the tumor replaces the entire thymic gland and invades into and through the pleura, pericardium, nerves, Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 233

3 Table 2. Ewing, Lymphosarcoma or thymoma Carcinoma arising from reticulum cells Spindle cell or myxosarcoma Lowenhaupt, I. Carcinoma of primitive epithelial reticulum II. Carcinoma of variegated cell pattern (carcinoma of early Hassall corpuscles) III. Carcinoma of granulomatous pattern (carcinoma of late Hassall corpuscles) IV. Carcinoma of thymic round cell: lymphoepithelioma V. Encapsulated thymoma VI. Carcinoma of adamantinomatous pattern Bernatz et al, Thymoma of predominantly Spindle cell type Lymphocytic type Mixed type (reticuloepithelial-lymphocytic) Epithelial type Levine and Rosai, Encapsulated thymoma Malignant thymoma I. Locally invasive thymoma Thymoma with lymphatic or hematogenous spread II. Cytologically malignant (thymic carcinoma) Marino and Müller-Hermelink, Medullary thymoma (medullary epithelial cells and scant, usually negligible lymphoid cells) Mixed thymoma (cortical and medullary epithelial cells, high number of lymphocytes) Predominantly cortical Predominantly medullary Cortical thymoma (cortical epithelial cells) Thymic carcinoma Kirchner et al, Medullary thymoma Mixed thymoma Predominantly cortical thymoma Cortical thymoma Well-differentiated thymic carcinoma Thymic carcinoma Suster and Moran, Thymoma Atypical thymoma Thymic carcinoma WHO, , 2004, 2015 WHO Type A Atypical type A variant a AB B1 B2 B3 C/thymic carcinoma b Selected Histomorphologic Classifications of Thymic Epithelial Tumors Well differentiated Moderately differentiated Poorly differentiated Morphologic Features Bland spindle/oval tumor cells, rare or no thymocytes Type A criteria and atypia (hypercellularity, increased mitotic count, focal tumor necrosis) Type A and type B1 or type B2 Scattered neoplastic cells in a background of thymocytes Presence of medullary elements Mixture of neoplastic cells and thymocytes Increased cytologic atypia of neoplastic cells Polygonal neoplastic cells with conspicuous nucleoli Thymic carcinoma Cytologic atypia Architecture distorted Desmoplasia Abbreviation: WHO, World Health Organization. a 2015 classification only. b Type C (1999 classification) was designated as thymic carcinoma in the 2004 and 2015 WHO classifications. and/or blood vessels. In addition, he described the discrete nodules in the pleura and pericardium that are not connected with the primary tumor as implants rather than true metastases. However, although Castleman acknowledged lymph node metastases, he avoided terms such as malignant thymoma or carcinoma of the thymic gland because (1) he did not observe any thymomas with embolic metastases and (2) he noted that encapsulated thymomas were morphologically similar to thymomas that spread within the mediastinum. In his opinion, tumors that were 234 Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

4 Table 3. Cytologic Characteristics of Epithelial Cells of the Benign Thymic Gland 22 Thymic Epithelial Cell Medullary Cortical Shape Spindle Stellate Nucleus Oval-spindle Oval-round Size Small-medium Medium-large Nucleolus þ/, not prominent Prominent Cytoplasm if present Scant, dense eosinophilic Scant, clear or faintly eosinophilic Abbreviation: þ/, nucleolus might or might not be present and if a nucleolus is present then it is not prominent. previously classified as malignant thymoma with metastases to distant organs did not have typical morphologic features of thymomas but were usually lymphomas, teratomas, or primary tumors of the lung, pancreas, or stomach that presented with large metastases to mediastinal lymph nodes. Moreover, he suggested that although tumors with implants and/or extension, invasion, or penetration of neighboring structures might be considered malignant, surgical removal of these tumors before implants have occurred or before the tumor extended within the mediastinum might have cured the patients. Recurrences, he felt, occurred likely because of incomplete resection. In 1961, with the hope that some prognostic significance could be gained through study of cell types, Bernatz et al 9 classified thymomas based on the predominant cell type (Table 2). Bernatz et al 9 reported on 138 patients who underwent surgery for thymoma at the Mayo Clinic Rochester. Based on the proposed morphologic classification, they found that patients with predominant lymphocytic or spindle cell thymomas have a better prognosis than patients with mixed cell type or predominant epithelial cell thymoma. Furthermore, predominant lymphocytic or spindle cell thymomas were more often encapsulated than thymomas with other predominant cell types. Moreover, based on the data from this study, Bernatz et al 9 concluded that lack of invasion is of more prognostic significance than cell type, although statistical analysis was not performed at that time. They realized, however, that the classification of thymomas would likely need to be further developed and concluded their article with the words from Effler and McCormack 15 that the intriguing subject of thymic neoplasms still requires new thought and clarification. The study by Bernatz et al 9 also showed the previously disputed benefit of thymectomy for at least some patients with myasthenia gravis. Eighteen of 64 patients (28.1%) with thymoma in the setting of myasthenia gravis who underwent thymectomy were well either without (n ¼ 11) or with (n ¼ 7) medication after a mean follow-up of 8 years. In a subsequent study, Bernatz et al 16 applied statistical analysis to outcome data of 80 patients with thymoma and myasthenia gravis (64 of which were also included in the original study of Bernatz et al 9 from 1961). The authors confirmed that myasthenia gravis did not affect survival of invasive thymomas. However, in noninvasive thymomas the survival of patients with myasthenia gravis was worse than for patients without that disease. Wilkins and Castleman 17 in 1979 did not find myasthenia gravis to be an adverse prognostic factor (anymore), possibly because of improved treatment. This was also confirmed in 1981 by Masaoka et al, 18 who showed that the 5-year survival of patients with thymoma and myasthenia gravis was similar to patients with thymoma without myasthenia gravis. However, the 10- year survival of patients with myasthenia gravis was worse in that study, which might have been because many deaths due to myasthenia gravis occurred during the early years of the study and, therefore, influenced the 10-year survival. A recent analysis of the retrospective Japanese database of TETs that included 598 TETs in patients with myasthenia gravis did not reveal any difference in 5- and 10-year overall or recurrence-free survival. 19 A recent multicenter study from Italy including 375 patients with thymoma showed a slight protective effect of the presence of myasthenia gravis on overall survival, although that was not confirmed in multivariate analysis. 20 These findings suggest that today the outcome of patients with thymoma and myasthenia gravis is at least as good as the outcome of patients with thymoma without myasthenia gravis. In 1976, Rosai and Levine 13 defined thymoma simply as an epithelial tumor originating from the thymic gland. The authors proposed to restrict the designation of thymoma to neoplasms of thymic epithelial cells regardless of the proportion of accompanying lymphocytes. The authors suggested that terms such as seminomatous thymoma, round cell thymoma, orgranulomatous thymoma should not be used anymore; these tumors should be designated according to the cell of origin, that is, germinoma (seminoma) of thymus, thymic carcinoid, orhodgkin disease, respectively. Rosai and Levine supported their belief that once the term thymoma is restricted to the tumor of epithelial thymic cells, with or without a lymphocytic component, all further subdivisions are artificial based on the review of 164 cases of thymoma. In a subsequent review, Levine and Rosai 21 added a component of thymic carcinoma to the classification (Table 2). Subsequently, TETs were classified in more detail and classifications focused on the relationship between neoplastic cells and their possible origin from the cortex and/or medulla of the benign thymic gland. Evidence had shown that in the benign thymic gland medullary epithelial cells are distinct from cortical epithelial cells based on morphology, ultrastructure, and immunology and therefore provide different microenvironments. 22 Furthermore, it was suggested that differences in nuclear and cytoplasmic features allowed the distinction between medullary and cortical thymic epithelial cells by microscopy (Table 3; Figure 1, A and B). Therefore, a classification into medullary predominant, cortical predominant, and mixed cell types was appealing. In 1985, Marino and Mueller-Hermelink 22 first proposed a classification of TET based on the compartment of the normal thymic gland from which the neoplastic epithelial cells likely arose (Table 2). In addition, thymic carcinomas were defined by the presence of invasive growth and almost pure composition of epithelial cells with cytologic criteria of malignancy. They applied this classification to a series of 58 thymomas and 13 thymic carcinomas and found that cortical thymomas had a tendency to occur in younger patients than other thymomas. 22 Furthermore, myasthenia gravis was only identified in patients with cortical thymoma. The more aggressive nature of cortical thymomas was reflected by their tendency for invasive growth and their clinical behavior, with 14 of 25 cases (56%) being locally invasive and/or having intrathoracic (13; 52%) or extrathoracic (1; 4%) metastases. A single case recurred. In addition, the authors noted differences in the lymphoid component, with medium- and large-sized lymphocytes more commonly seen in pure or predominantly Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 235

5 Figure 1. Epithelial cells in the benign thymic gland. The benign thymic gland contains large, stellate cortical epithelial cells that are characterized by round or oval medium-to-large nuclei with prominent nucleoli and scant eosinophilic cytoplasm (A, arrows) and spindle medullary epithelial cells with oval-to-spindle small-to-medium nuclei without or with inconspicuous nucleoli and scant eosinophilic cytoplasm (B, arrow). The epithelial cells are highlighted by keratin AE1/AE3 immunostain (A and B, insets) (hematoxylin-eosin, original magnification 3600 [A and B]; keratin AE1/AE3 immunostain, original magnification 3600 [A and B, insets]). cortical thymoma whereas small pleomorphic lymphocytes were mostly seen in medullary or mixed thymoma with medullary predominance. Moreover, the number of lymphocytes was high in cortical and mixed thymomas, whereas pure medullary thymomas had only rare lymphocytes. The concept by Marino and Müller-Hermelink 22 of classifying TET according to the structural components of the normal thymus was further developed by Kirchner et al 23 in 1989 based on a study of 95 TETs, patient s outcome, and proliferation assays of 12 TETs (Table 2). Kirchner et al 23 introduced a category of well-differentiated thymic carcinoma, defined as TET with incomplete loss of organotypic differentiation, tightly packed epithelial cells with slight to moderate atypia, and some immature CD1-positive T cells. Small areas of cortical differentiation could be present in well-differentiated thymic carcinoma. In contrast to welldifferentiated thymic carcinoma, other thymic carcinomas were defined by lack of organotypic features. Based on outcome studies, Kirchner et al 23 considered medullary and mixed thymomas as benign; predominantly cortical thymomas, cortical thymomas, and well-differentiated thymic carcinomas as low-grade malignant tumors with increasing invasiveness and metastatic capacity; and thymic carcinoma as overtly malignant tumors. In vitro studies confirmed that the proliferative rate of neoplastic epithelial cells correlates with the different tumor types and their growth behavior in vivo. For instance, cells from mixed thymomas (n¼3) did not grow in vitro or ceased to proliferate after less than 3 passages, whereas cells from well-differentiated thymic carcinomas (n ¼ 3) had the highest growth rate among the Table 4. Organotypic Features of Differentiation of the Thymus as Defined by Suster and Moran 24 Lobular architecture Dual cell population (neoplastic thymic epithelial cells, thymic lymphocytes) Dilated perivascular spaces Areas of medullary differentiation (þ/ Hassall corpuscles) a a þ/, may or may not be present. TETs tested, could be passaged for a maximum of 35 times, and were kept in culture for up to 8 months. The proliferative rate of epithelial cells of cortical (n ¼ 3) and predominantly cortical thymomas (n ¼ 3) was between that of mixed thymomas and well-differentiated thymic carcinomas. In 1999, Suster and Moran 24,25 proposed a 3-tiered classification that was based upon morphologic features of differentiation (Table 2). This classification subtyped TETs according to their degree of cytologic atypia, presence of organotypic features of thymic differentiation, and closeness to benign thymus. Organotypic features of differentiation were defined as features that most closely resemble the normal appearance of the thymus in its mature or involuted state and are summarized in Table 4. Areas of medullary differentiation were described as well-circumscribed foci containing plump epithelial cells with scant lymphocytes within an otherwise cortical-appearing neoplastic population of cells, with or without Hassall corpuscles. According to this classification, thymomas exhibit all or most organotypic features, thymic carcinomas do not have any or have only minimal organotypic features, and atypical thymomas show intermediate morphologic features. The classification is illustrated in Figure 2, A through D. In addition, the authors considered all thymomas to potentially be malignant. In 1989, a WHO [World Health Organization] committee for the histologic classification of tumours of the thymus was organized by Dr Juan Rosai. Subsequently, pathologists from 8 countries debated during 1 decade whether a histologic subclassification of thymomas is possible and useful. Although some pathologists favored a classification that just distinguished noninvasive from invasive thymomas, others thought that histologic types of thymoma correlated with their aggressiveness and clinical behavior and therefore further morphologic subclassification would be useful. Subsequently, a compromised histologic classification was published. 26 Because the association between thymoma subtype and compartment of normal thymic gland was still controversial, the terminology chosen by the WHO was noncommittal and was designated by 236 Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

6 Figure 2. Histomorphologic classification by Suster and Moran. 24 A, The well-differentiated thymoma is characterized by preserved organotypic features including lobulated growth pattern and a mixture of epithelial tumor cells and thymocytes (inset). B, The moderately differentiated atypical thymoma shows a lobulated architecture; however, the neoplastic epithelial cells are cytologically more atypical and at most only occasional thymocytes are scattered throughout the tumor (inset). C and D, The poorly differentiated thymic epithelial tumor, the thymic carcinoma, lacks a lobulated architecture and is characterized by irregular tumor cell nests surrounded by a desmoplastic stromal reaction (C). The tumor cells are cytologically atypical and thymocytes are absent (D) (hematoxylin-eosin, original magnifications [A and B], 320 [C], and 3400 [A and B, insets, and D]). letters and numbers (Table 2). All thymomas composed of bland-appearing spindle and/or oval cells were classified as type A; thymomas that contained dendritic or plump ( epithelioid ) cells were categorized as type B. Type B thymomas were further subdivided based on the proportional increase in tumor cells in relation to thymocytes, increasing cytologic atypia, and presence or absence of medullary differentiation. Figure 3, A through F, illustrates the morphologic features of the WHO classification. In addition to these classic thymomas, uncommon thymomas were described, including micronodular thymoma (Figure 4, A), microscopic thymoma (Figure 4, B), and metaplastic thymoma (Figure 4, C and D). Thymic neuroendocrine tumors were defined identically to neuroendocrine tumors in the lung, including typical carcinoid tumor, atypical carcinoid tumor (defined as 2 10 mitoses per 10 high-power fields and/or necrosis), large cell neuroendocrine carcinoma, and small cell carcinoma. The 1999 WHO classification 26 emphasized that the classification of cytoarchitectural features of thymoma should occur independent of staging because the classification based on invasive/metastasizing properties of the tumour relates more closely to recurrence and outcome than the one based on cytoarchitectural features. The WHO classification was revised in 2004 to include descriptions of clinical symptoms, macroscopic findings, immunohistochemical characteristics, genetic features, and prognostic data. 27 Type C thymoma (1999 WHO classification 26 ) was now defined as thymic carcinoma; the morphologic subtypes of thymomas remained unchanged. The most recent WHO classification, 28 published in 2015, advocated a similar histomorphologic classification that had retained the designation of the tumors by letters and numbers. The 2015 WHO classification used an interdisciplinary approach to the diagnosis of TET and included contributions from radiologists, thoracic surgeons, and oncologists. Epidemiologic and prognostic data for the WHO classification were derived, in part, from the worldwide retrospective database of the International Thymic Malignancy Interest Group (ITMIG), which comprises more than 6000 cases. Findings on imaging studies, Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 237

7 Figure 3. Histomorphologic classification by the World Health Organization. A, Type A thymomas are characterized by bland-appearing spindle cells with absent or only scattered thymocytes (inset). B, This type AB thymoma comprises a type B2 component (upper left-hand side, upper inset) and a type A component (lower right-hand side, lower inset), which are next to each other in this case. C, At low magnification a type B1 thymoma is in general recognized by medullary islands (arrows) that sometimes contain Hassall corpuscle like particles (upper inset). There are only scattered neoplastic epithelial cells in a background of predominantly thymocytes (lower inset). D, On low magnification type B2 thymomas often have a starry-sky appearance because of the mixture of medium-to-large neoplastic epithelial cells and thymocytes (inset). E, Type B3 thymomas have a lobulated appearance on low power; thymocytes are only scattered or absent (inset). F, The thymic carcinoma has a distorted architecture and is composed of irregular nests of cytologically atypical tumor cells (inset) surrounded by a desmoplastic stroma (hematoxylin-eosin, original magnifications 340 [A, B, and D through F], 320 [C], and 3600 [A through F, insets]). specifically computer tomography/positron emission tomography and cytologic features, were also included. Some histomorphologic features and immunohistochemical criteria were refined in an attempt to enhance the reproducibility of subtyping of thymomas and to facilitate the distinction between thymomas and thymic carcinomas. 29 For instance, to help with the distinction between type B1 and B2 thymoma, the definition of type B1 thymomas included features such as thymuslike architecture and cytology, abundance of immature T cells, areas of medullary differentiation (medullary islands), and paucity of polygonal or dendritic epithelial cells without clustering (,3 contiguous epithelial cells) (Figure 5, A and B). Hassall corpuscles or perivascular spaces were considered optional. In addition, the distinction between type A and type AB thymomas was discussed in more detail. Although both types contain bland-appearing spindle/oval cells, they should be distinguished from each other by a low ( easily countable, type A) or high (TdT-positive, T-cells impossible to count, and/ or moderate infiltrate of TdTþ lymphocytes [difficult to count] in. 10% of the tumour areas, type AB) content of lymphocytes. Furthermore, an atypical type A thymoma variant was introduced. This TET is characterized by features of conventional type A thymoma with bland-appearing spindle/oval cells and atypical findings including hypercellularity, increased mitotic activity, and focal necrosis. This variant was included in the WHO classification because studies have shown an association between necrosis in type A thymomas and recurrence and extrathoracic metastasis. 30 Moreover, in a study of type A and AB thymomas, necrosis was associated with stage in univariate and multivariate analysis. 31 However, neither study showed an association between mitotic activity and stage or outcome in type A 30 or type A and AB thymomas. 31 The 2015 WHO classification recommended reporting of all thymoma subtypes in 10% increments (except type AB thymomas) if more than one subtype is identified in a resection specimen. Furthermore, all subtypes of thymoma were considered malignant because they can show an aggressive behavior. Exceptions were micronodular and microscopic thymoma, in which no fatal outcome has been reported. It was also suggested that immunohistochemistry might be used for the diagnosis and Table 5. Thymic Carcinoma Subtypes According to 2015 World Health Organization Classification 28 Squamous cell carcinoma Basaloid carcinoma Mucoepidermoid carcinoma Lymphoepithelioma-like carcinoma Clear cell carcinoma Sarcomatoid carcinoma Adenocarcinomas Thymic carcinoma NUT carcinoma Undifferentiated carcinoma Others Thymic neuroendocrine tumors including: Carcinoid tumors (typical, atypical carcinoid) Large cell neuroendocrine carcinoma Small cell carcinoma Abbreviation: NUT, nuclear protein of testis. 238 Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

8 Figure 4. Uncommon thymomas. A, The micronodular thymoma with lymphoid stroma is characterized by nests and strands of bland-appearing spindle tumor cells (inset) in a lymphocytic background. The background might contain germinal centers (arrow). B, This microscopic thymoma measures 0.9 mm in greatest dimension and lacks a distinct tumor capsule. It is composed of bland-appearing spindle cells (inset). C and D, A metaplastic thymoma is composed of nests and strands of bland-appearing tumor epithelial cells, some of which can contain nuclear pseudoinclusions (D, right-hand side) and are surrounded by metaplastic spindle cells (D, left-hand side) (hematoxylin-eosin, original magnifications 320 [A and C], 340 [B], and 3400 [A and B, insets, and D]). Figure 5. Distinction of type B1 from type B2 thymoma using the World Health Organization 2015 criteria. A type B1 thymoma (A) is defined by scattered epithelial tumor cells with fewer than 3 clustering together (arrows), in contrast to a type B2 thymoma (B), in which 3 or more tumor cells might cluster (arrows) (hematoxylin-eosin, original magnification 3600). Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 239

9 Table 6. Selected Proposed Staging Systems for Thymic Epithelial Tumors 1961 Through 1994 Bernatz et al, Noninvasive Invasive Bergh et al, Stage Definition I Intact capsule; growth within capsule II Pericapsular growth into mediastinal fat tissue III Invasive growth into surrounding organs, intrathoracic metastases or both Wilkins and Castleman, Stage Definition I Intact capsule or growth within capsule II Pericapsular growth into the mediastinal fat tissue or adjacent pleura or pericardium III Invasive growth into the surrounding organs, intrathoracic metastases, or both Masaoka et al, , and modified Masaoka (Koga et al, ) staging Stage Masaoka Modified Masaoka (Koga) I Macroscopically completely encapsulated, microscopically no capsular invasion Macroscopically completely encapsulated, microscopically no capsular invasion II 2. Microscopic invasion into capsule A. Microscopic transcapsular invasion 1. Macroscopic invasion into surrounding fat or mediastinal pleura B. Macroscopic invasion into extracapsular soft tissue, or tumor grossly adherent to mediastinal pleura or pericardium III Macroscopic invasion into neighboring organs Macroscopic invasion into neighboring organs IV A. Pleural or pericardial dissemination B. Lymphogenous or hematogenous metastasis Staging Classification of the Groupe d Étude des Tumeurs Thymiques, Stage Definition IA Encapsulated tumor, totally resected IB Macroscopically encapsulated tumor, totally resected, perioperative suspicion of mediastinal adhesion or potential capsular invasion II IIIA IIIB IVA IVB Invasive tumor, totally resected Invasive tumor, subtotally resected Invasive tumor, biopsy Supraclavicular metastasis or distant pleural droplets Distant metastasis A. Pleural or pericardial dissemination B. Lymphogenous or hematogenous metastasis TNM classification of thymic epithelial tumors by Yamakawa et al, ; pathologic TNM classification of thymic epithelial tumors by Tsuchiya et al, ; and TNM classification of malignant thymic epithelial tumors as recommended by the 2015 World Health Organization 28 classification Yamakawa et al Tsuchiya et al World Health Organization 2015 TX Primary tumor cannot be assessed T0 T1 T2 Macroscopically completely encapsulated and microscopically no capsular invasion Macroscopically adhesion or invasion into surrounding fatty tissue or mediastinal pleura, or microscopic invasion into capsules Macroscopically completely encapsulated and microscopically no capsular invasion Tumor breaking through capsule, invading thymus or fatty tissue (maybe adherent to mediastinal pleura but not invading neighboring organs) No evidence of primary tumor Macroscopically completely encapsulated and microscopically no capsular invasion Tumor invades pericapsular connective tissue 240 Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

10 Table 6. Continued T3 Invasion into neighboring organs, such as pericardium, great vessels, and lung Breaking through the mediastinal pleura or pericardium, or invading neighboring organs, such as great vessels and lung Tumor invades into neighboring structures, such as pericardium, mediastinal pleura, thoracic wall, great vessels, and lung T4 Pleural or pericardial dissemination Pleural or pericardial dissemination Pleural or pericardial dissemination NX Regional lymph nodes cannot be assessed N0 No lymph node metastasis No lymph node metastasis No lymph node metastasis N1 Metastasis to anterior mediastinal Metastasis to anterior mediastinal Metastasis to anterior mediastinal N2 lymph nodes Metastasis to intrathoracic lymph nodes except anterior mediastinal lymph nodes lymph nodes Metastasis to intrathoracic lymph nodes except anterior mediastinal lymph nodes lymph nodes Metastasis to intrathoracic lymph nodes except anterior mediastinal lymph nodes N3 Metastasis to extrathoracic lymph nodes Metastasis to extrathoracic lymph nodes M0 No hematogenous metastasis No hematogenous metastasis No distant metastasis M1 Hematogenous metastasis Hematogenous metastasis Distant metastasis Stage I T1 N0 M0 T1,2 N0 M0 T1 N0 M0 II T2 N0 M0 T1,2 N1 M0 T2 N0 M0 III T3 N0 M0 T3 N0,1 M0 T1,2 N1 M0 T3 N0,1 M0 IV T4 N any M0 T any N2,3 M0 T any N any M1 IVa T4 N0 M0 T4 N0,1 M0 IVb T any N1,2 or 3 M0 T any N2,3 M0 T any N any M1 IVc T any N any M1 Metastases in scalene and/or supraclavicular lymph nodes subtyping of thymomas with ambiguous histology. Thymic carcinoma subtypes have been broadened and now include the NUT carcinoma (Table 5). EVOLUTION OF THE STAGING CLASSIFICATION OF THYMIC EPITHELIAL TUMORS Staging classifications of TET in general are based upon invasion, implants, lymph node involvement, and/or distant metastases. During the past half-century, at least 14 staging systems have been proposed, even though many of these were not outright designated as staging systems (Table 1). This review will focus on those that introduced new and interesting concepts of classification. In 1961, Bernatz et al 9 divided thymomas into noninvasive and invasive thymomas (Table 6). 9 This categorization was based on their study that showed that only 6 of 27 patients (22.2%) with invasive thymoma lived 5 or more years following surgery, in contrast to 49 of 64 patients (76.5%) with noninvasive thymoma, suggesting that patients with invasive thymoma had worse survival than patients with noninvasive thymoma. Similarly, in 1976, Rosai and Levine 13 suggested that thymomas need only be separated into encapsulated versus invasive tumors for predicting outcome and guiding therapy. Rosai and Levine found that well-encapsulated thymomas without myasthenia gravis remain stable and surgical excision is curative, even though the patient might develop a paraneoplastic syndrome or local recurrence, which can occur either as multiple pleural implants or as localized mediastinal tumor. In contrast, invasive thymomas, although histologically indistinguishable from encapsulated thymoma, have a worse outcome. In 1978, Bergh et al 32 developed a 3-tiered staging system that further subclassified invasive thymomas (Table 6). When applied to 43 thymomas, that staging system revealed considerable differences in survival between stage II and III thymomas, supporting that the extent of invasion has prognostic value. Wilkins and Castleman 17 modified the classification by Bergh et al slightly in 1979, adding invasion into pleura and pericardium to stage II (Table 6). The authors confirmed that patients with invasive thymomas had an overall worse outcome than patients with noninvasive thymomas; however, the authors did not reach any conclusions in regards to outcome of patients with stage II versus stage III thymoma. In 1981, Masaoka et al 18 pioneered today s most commonly used staging system for thymomas (Table 6). This system was developed for clinical staging of thymomas assuming that all thymomas have a variable degree of malignant potential and prognosis of thymoma is determined from its stage, which is based on invasiveness of the tumor through the capsule into surrounding tissue and organs and metastases. This classification was designed upon their observation that thymomas initially grow locally, subsequently infiltrate or disseminate, and finally metastasize. It was sought to guide therapy, to evaluate the results of surgery, and to define the prognosis of a patient. When this staging system was applied to 96 thymomas, they showed that survival differed based on stage, with 5-year survivals of 92.6% (stage I), 85.7% (stage II), 69.6% (stage III), and 50% (stage IV). In 1994, Koga et al 33 modified the Masaoka staging, changing invasion into the tumor capsule (Masaoka stage II/2) to microscopic invasion through the capsule (transcapsular; Koga stage IIA) to accommodate the most common point of view of pathologists that a tumor is invasive if it grows through the tumor capsule (Table 6). Therefore, thymomas invading into but not through the tumor capsule were now classified as stage I in the Koga staging system. It was confirmed later by Roden et al 4 that the prognosis is similar for patients with thymomas infiltrating into but not through the tumor capsule when compared with patients with encapsulated thymomas. Applying this system to 79 thymomas, Koga et al found Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 241

11 that although the outcome did not differ significantly between stages I and II, there were significant differences between stages II and III and stages II and IV. The Koga staging system is now often referred to as the modified Masaoka or the Masaoka-Koga staging system. This staging system is probably worldwide the most commonly used staging system for thymomas today and is recommended by the WHO. 29 In 1982, the French Groupe d Étude des Tumeurs Thymiques proposed a clinical classification that encompassed not only extent of invasion and metastases but also completeness of resection (Table 6). 34,35 In 1991, Yamakawa and colleagues 36 proposed a tentative tumor-node-metastasis (TNM) classification that was similar to the Masaoka staging system but distinguished between lymphogenous and hematogenous metastases (Table 6). Lymphogenous metastases occurred in a few patients and were thought to progress from anterior mediastinal lymph nodes to intrathoracic and later to extrathoracic lymph nodes. Hematogenous metastases did not reveal any particular characteristics. The T stage was adopted from the Masaoka classification. In contrast to TNM classifications of other organs, the T stage for TET did not consider size of the tumor because it was thought that the malignant potential of thymoma was due to its invasiveness and its tendency to disseminate rather than its size. The TNM classification was applied to 207 thymomas. When evaluating stage IVB cases (thymomas with lymphogenous and/or hematogenous metastases), no association between stage and prognosis or treatment was identified, possibly because of the small number of cases (8 of 207; 3.9%). The classification was also applied to 13 thymic carcinomas and 6 thymic carcinoid tumors. In this population, there was a much higher frequency of stage IVB cases, and the TNM classification was considered to be more useful in these tumors. Tsuchiya et al 37 thought that a pathologic TNM staging system would be applicable to thymic carcinomas and thymic carcinoid tumors but not to thymomas, as Yamakawa et al 36 had suggested, and proposed a modification to the TNM staging system in 1994 (Table 6). Stage pt2 now included thymic carcinomas with invasion through the tumor capsule; tumors that invaded through the pleura or the pericardium were classified as stage pt3. The staging groups were also modified. Tumors with N1 disease were now distributed into stages II to IV based upon the T stage because N1 disease was considered resectable and curable. The authors validated the modified TNM staging system on 16 thymic carcinomas. Although there were no stage II thymic carcinomas in this study, survival curves separated them between stages I and III or IV and stages III and IV; the differences were not statistically significant, likely again because of the small number of cases. Moreover, this study predominantly included squamous cell carcinomas and 2 undifferentiated and 1 basaloid carcinoma, and lacked other carcinoma types of high-grade histology such as small cell carcinoma, lymphoepithelioma-like carcinoma, sarcomatoid carcinoma, and clear cell carcinoma. Carcinoid tumors were also not evaluated. In 1997, Suster and Moran 25 developed a simple 3-tiered staging classification that was proposed to be usable to stage both thymic carcinomas and thymomas (Table 7). Weissferdt and Moran 38 proposed a 3-tiered staging classification for thymic carcinomas in 2012 (Table 7) with the purpose of simplifying the existing TNM classification. This classifica- Table 7. Selected Proposed Staging Systems for Thymic Epithelial Tumors From 1997 Through 2014 Suster and Moran, Stage Definition I Well encapsulated tumor II Infiltrative or locally invasive tumor III Lymph node or distant metastasis Staging of Thymic Carcinomas as Proposed by Weissferdt and Moran, Definition T1 Tumor limited to thymic gland T2 Tumor invading visceral pleura, lung, pericardium, great vessels, chest wall or diaphragm T3 Direct extrathoracic tumor extension, beyond thoracic inlet or diaphragm N0 No lymph node metastasis N1 Lymph node metastasis to intrathoracic lymph node M0 No distant metastasis M1 Distant metastasis Stage I T1 N0 M0 II T2 N0 M0 III T3 N0 M0 T any N1 M0 T any N any M1 Staging of Thymomas as Proposed by Moran et al, Stage Definition 0 Encapsulated tumor I Tumor invasive into perithymic adipose tissue II Direct invasion A Innominate vein, mediastinal pleura, lung B Pericardium C Great vessels (aorta, superior vena cava), heart III Metastatic disease A Intrathoracic structures, diaphragm, lymph nodes B Extrathoracic invasion Staging of Thymic Epithelial Tumors as Proposed by IASLC/ ITMIG, Definition (Involvement of) a T1 a Encapsulated or unencapsulated, with or without extension into mediastinal fat b Extension into mediastinal pleura T2 T3 T4 N0 N1 N2 M0 M1 a b Pericardium Lung, brachiocephalic vein, superior vena cava, chest wall, phrenic nerve, hilar (extrapericardial), pulmonary vessels Aorta, arch vessels, main pulmonary artery, myocardium, trachea, esophagus No lymph node involvement Anterior (perithymic) lymph nodes Deep intrathoracic or cervical lymph nodes No metastatic pleural, pericardial, or distant sites Separate pleural or pericardial nodule(s) Pulmonary intraparenchymal nodule or distant organ metastasis Stage I T1 N0 M0 II T2 N0 M0 IIIa T3 N0 M0 IIIb T4 N0 M0 IVa T any N1 M0 T any N0 or 1 M1a IVb T any N2 M0 or 1a T any N any M1b Abbreviations: IASLC, International Association of Study of Lung Cancer; ITMIG, International Thymic Malignancy Interest Group. a A tumor is classified according to the highest level of invasion regardless whether invasion of structures of lower levels is present or not. 242 Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden

12 Table and 2004 World Health Organization Criteria for Invasion and Metastasis 26,27 World Health Organization Category Encapsulated Minimally invasive Widely invasive Implants Lymph node metastasis Distant metastases Definition No invasion through the tumor capsule Invasion through tumor capsule into surrounding adipose tissue Invasion into neighboring organs including lung, pericardium, great vessels Tumor nodules on pleura, pericardium, and diaphragm Mediastinal and supraclavicular lymph nodes Lung, liver, skeletal system, others tion was developed based on outcome data of 33 patients with thymic carcinoma. At the same time, Moran et al 39 recommended a staging solely for thymomas (Table 7). The authors introduced a stage 0 or in situ malignancy, which most likely could be controlled by complete resection alone. Only invasive tumors were assigned a stage. However, the authors acknowledged that recurrence has been reported even in encapsulated thymomas. When the authors applied this staging system to 250 patients with thymoma, significant differences in overall and recurrence-free survival were identified when comparing stages 0 and I with stages II and III; no differences were found in outcome between stages 0 and I. Furthermore, there were no differences in outcome between stages IIa, b, and c. The authors considered, however, that distinction was still important because of possible differences in treatment. The prognostic significance of the proposed Moran staging for thymomas was later confirmed by Roden et al, 40 although in that study, patients with stage 0 disease had a significantly better overall survival than stage I patients. Currently, neither the American Joint Committee on Cancer nor the Union for International Cancer Control provides a staging system for thymoma or thymic carcinoma. The modified Masaoka staging system (Masaoka-Koga staging system) appears to be the most commonly used staging system for thymomas, whereas the TNM staging as recommended by the WHO (Table 6) might be used for thymic carcinoma. 28,41 To facilitate staging, the 1999 and 2004 WHO included morphologic criteria to facilitate the evaluation of invasion and metastases (Table 8; Figure 6, A through F). 26,27 There is a need for a staging system that can be applied to all TETs, including thymomas, thymic carcinomas, and thymic neuroendocrine tumors. 42 Therefore, the International Association for the Study of Lung Cancer (IASLC) together with ITMIG proposed a staging system in 2014 for the forthcoming (eighth) edition of the TNM classification of malignant tumors (Table 7). 42 This classification requires microscopic evidence of findings for pathologic staging. This staging system was validated using a worldwide retrospective database led by ITMIG and supplemented by cases from the Japanese Association for Research on the Thymus and Figure 6. Pathologic criteria (2004 World Health Organization) to facilitate staging according to the modified Masaoka (Masaoka-Koga) staging system. A, Encapsulated thymoma, stage I. In some thymomas, the tumor capsule might be thin or the tumor might invade into the capsule (arrow and inset); however, no invasion of the tumor through the capsule is identified. B, Minimally invasive thymoma, stage IIA. The thymoma has invaded through the tumor capsule into the surrounding adipose tissue (arrow and inset). C, Widely invasive thymoma, stage IIIA. This thymoma invades directly into lung parenchyma. D through F, Metastatic thymoma, stage IVB. Thymoma can metastasize to various organs, including the lung (D, type A thymoma; D inset), ovary (E; arrow points toward ovarian stroma) or bone (F; F inset shows metastatic thymic carcinoma on the left and a megakaryocyte of the bone marrow on the right with arrow pointing toward a megakaryocyte) (hematoxylin-eosin, original magnifications [A through D], 3100 [A, inset], 3200 [B, inset], 3400 [D and F, insets], and 340 [E and F]). Arch Pathol Lab Med Vol 141, February 2017 Classification of Thymic Epithelial Tumors Roden 243