Perspectives on testicular germ cell neoplasms

Liang Cheng MD, Bingjian Lyu MD, Lawrence M. Roth MD

PII: S0046-8177(16)30193-9
DOI: doi: 10.1016/j.humpath.2016.08.002
Reference: YHUPA 3980

To appear in: Human Pathology

Received date: 19 May 2016

Revised date: 9 August 2016
Accepted date: 11 August 2016

Please cite this article as: Cheng Liang, Lyu Bingjian, Roth Lawrence M.,
Perspectives on testicular germ cell neoplasms, Human Pathology (2016), doi:
10.1016/j.humpath.2016.08.002

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 ACCEPTED MANUSCRIPT

Revised Manuscript YHUPA-D-16-00343R2

Perspectives on testicular germ cell neoplasms

Liang Cheng, MD1,2, Bingjian Lyu, MD3, Lawrence M. Roth, MD1

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From the Department of Pathology and Laboratory Medicine1, and Urology2, Indiana

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University School of Medicine, Indianapolis, IN 46202, USA; Department of Surgical

Pathology3, Womens Hospital, School of Medicine, Zhejiang University, China, 310003

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Running Head: Perspectives on testicular germ cell tumors
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Total number of text pages, 36; number of tables, 2; number of figures, 10.

Keywords: Testis; germ cell tumors; teratoma; tumorigenesis; histogenesis; differential

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diagnosis.
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Conflicts of interest: None

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Funding: This research did not receive any specific grant from funding agencies in the

public, commercial, or not-for-profit sectors.

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Address correspondence and reprint requests to Liang Cheng, MD, Department of

Pathology and Laboratory Medicine, Indiana University School of Medicine, 350 West

11th Street, IUHPL Room 4010, Indianapolis, IN 46202, USA. Telephone: 317-491-

6442; Fax: 317-491-6419; E-mail: liang_cheng@yahoo.com

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Key points/Highlights

Germ cell neoplasms show significant differences in frequency, histological

appearance, and biological behavior depending on whether they occur in the testis

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or ovary. Both genetic and epigenetic factors likely account for these

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dissimilarities.

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Careful morphologic observations supported by detailed clinical information are

fundamental to the interpretation of perplexing gonadal germ cell tumors.

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Newly available antibodies and molecular diagnostic studies, when necessary, can

contribute to a more specific pathological interpretation in challenging cases.

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Effective integration of classical morphologic observations and emerging novel

molecular studies will likely result in optimal patient care; however, diagnostic
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pathology will continue to play an essential role in this process for the foreseeable
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future.
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Summary Our knowledge of testicular germ cell neoplasms has progressed in the last

few decades due to the description of germ cell neoplasia in situ (GCNIS) and a variety

of specific forms of intratubular germ cell neoplasia, the discovery of isochromosome

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12p and its importance in the development of invasiveness in germ cell tumors (GCTs),

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the identification of specific transcription factors for GCTs, and the recognition that a

teratomatous component in mixed GCT represents terminal differentiation.

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Isochromosome 12p and 12p overrepresentation, collectively referred to as 12p

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amplification, are fundamental abnormalities that account for many types of malignant

GCTs of the testis. Embryonal carcinoma is common in the testis but rare in the ovary,
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whereas the converse is true for mature cystic teratoma. Spermatocytic tumor occurs only

in the testis; it has not been described in the ovary or extragonadal sites. The origin of
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ovarian mature cystic teratoma is similar to that of prepubertal type testicular teratoma
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and dermoid cyst at any age in that it arises from a non-transformed germ cell, whereas

postpubertal type testicular teratoma arises from a malignant germ cell, most commonly
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through the intermediary of GCNIS. Somatic neoplasms, often referred to as monodermal

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teratomas, arise not infrequently from mature cystic teratoma of the ovary, whereas such

neoplasms are rare in testicular teratoma with the exception of carcinoid. Integration of

classical morphologic observations and emerging novel molecular studies will result in

better understanding of the pathogenesis of GCTs and will optimize patient therapy.

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1. INTRODUCTION

Germ cell tumors (GCTs) are the most common category of testicular neoplasms.

It is important to correctly diagnose the various types of GCT and to distinguish them

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from epithelial and sex cordstromal tumors so that optimal therapy and an accurate

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prognosis can be determined [1, 2]. In this article, we discuss the latest classification of

testicular GCTs in the recently published WHO Classification of Tumours of the Urinary

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System and Male Genital Organs [3]. The topics of tumors composed of germ cells and

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sex cord derivatives and disorders of sex development are beyond the scope of this

article.
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2. HISTOGENESIS AND PUTATIVE PRECURSORS OF GERM CELL TUMORS
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Both genetic and epigenetic factors are likely responsible for the myriad of
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differences between testicular and ovarian GCTs [4, 5]. Most fundamental is the

development of the testis and its neoplasms under the influence of the sex-determining
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region Y (SRY) gene on the short arm of the Y chromosome, whereas the ovary and its
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tumors develop without its influence [6]. Because of genomic imprinting, the paternal

and maternal sets of chromosomes have different functionality due to parental specific

epigenetic modification of the genome [4]. The genetic hallmark of the postpubertal types

of malignant GCT is chromosome 12p abnormalities, including isochromosome 12p and

chromosome 12p overrepresentation (Fig. 1) [7, 8].

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3. CLASSIFICATION OF GERM CELL NEOPLASMS

Our current classification of testicular GCTs modified from the most recent WHO

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Classification of Tumours the Urinary System and Male Genital Organs is shown in

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Table 1 [3]. One of the major features of the current classification of testicular GCTs is

the adoption of the division of teratoma and yolk sac tumor into prepubertal and

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postpubertal types; however, after this distinction was made, it was recognized that the

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prepubertal types could be detected in patients at a postpubertal age. On reflection, this

observation should not be considered surprising because it is not currently possible to

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determine the time of inception of a neoplasm. The age of the patient at the time of

detection presumably would be dependent on the age at inception of the neoplasm, its
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growth rate, its location in a cryptorchid or descended testis, and the frequency of
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examination by the individual, family, or clinician. None of these factors can be

controlled. Furthermore, postpubertal types can occur in prepubertal individuals who

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have disorders of sex development. Although it is well documented that prepubertal type
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neoplasms can present at an age beyond childhood, they nevertheless maintain discrete

characteristics that distinguish them from adult type testicular GCTs [9].

4. NONINVASIVE GERM CELL NEOPLASIA

In the recently published WHO Classification of Tumours of the Urinary System

and Male Genital Organs, the term germ cell neoplasia in situ (GCNIS) has replaced the

older terms intratubular germ cell neoplasia, unclassified and carcinoma in situ [3].

GCNIS has been recognized as a common precursor lesion for most malignant testicular

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GCTs since 1972 (Fig. 1). It is found in the adjacent seminiferous tubules of

approximately 88% of adult testicular GCTs and in approximately 5% of cases in the

contralateral testis [10-13]. However, GCNIS is absent in yolk sac tumor and teratoma of

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the prepubertal testis and in spermatocytic tumor of the adult testis.

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The specific forms of intratubular germ cell neoplasia include intratubular

seminoma and embryonal carcinoma, and, rarely, intratubular yolk sac tumor and

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teratoma (Fig. 2). Intratubular trophoblastic cells occur occasionally in association with

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seminoma; however, an intratubular component of choriocarcinoma or other

trophoblastic tumors has not been described. It is likely that the specific forms of
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intratubular germ cell neoplasia are derived either directly from transformation of

primordial germ cells or from intratubular differentiation of GCNIS [14].

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5. INVASIVE POSTPUBERTAL TYPE GERM CELL NEOPLASMS

5.1. Pure seminoma

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Testicular classical seminoma and ovarian dysgerminoma are morphologically

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identical and are collectively known as germinoma, particularly when they occur in

midline extragonadal locations including the sacrococcygeal area, retroperitoneum,

mediastinum, and central nervous system. Germinoma is arguably the most primitive of

GCTs, being derived from transformed primordial germ cells [15]. GCNIS is the

precursor of the great majority of classical seminomas as well as the other testicular

malignant GCTs. No corresponding precursor is recognized in the ovary although

gonadoblastoma is the precursor of germinoma in patients with disorders of sex

development. In its pure form (tumor with a single histologic type), seminoma is the most

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frequently encountered GCT of the testis. Seminomas are sensitive to chemotherapy and

radiotherapy; however, the latter is rarely employed at present because of greater

morbidity.

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Seminoma shows a characteristic pattern of monomorphic primitive germ cells

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separated by delicate fibrous tissue, variably infiltrated by lymphocytes, that divides the

neoplastic cells into lobules, cords, or columns (Fig. 3). Syncytiotrophoblasts occur in

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10-20% of cases but do not alter the prognosis. Granulomatous change occurs in about

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25% cases and sometimes is so extensive that it may mask the original features of the

neoplasm. In well-fixed specimens, neoplastic cells show the typical cytological features
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including round or polygonal shape, well-defined cytoplasmic borders, round or oval

nuclei with regular nuclear membranes, and obvious nucleoli. Morphological variants,
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including microcystic, cribriform, and solid tubular arrangements, can be encountered.

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Immunohistochemically, seminomas are positive for OCT4, SALL4, CD117 (c-

kit), podoplanin (D2-40), NANOG, SOX17, and PLAP and negative for CD30, GATA3,
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glypican-3, and SOX2 (Table 2) [16].

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5.2 Pure embryonal carcinoma

Embryonal carcinoma is a primitive GCT distinct from seminoma that is

relatively common in the testis but exceedingly rare in the ovary. Currently, testicular

embryonal carcinoma is thought to arise from GCNIS either through the intermediary of

intratubular embryonal carcinoma or from seminoma [14].

The most common architectural patterns are solid, glandular, and papillary (Fig.

4). Less common morphological variants including nested, micropapillary, anastomosing

and sieve-like, glandular, pseudopapillary, blastocyst-like, and appliqu patterns were

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described recently [17, 18]. Necrosis is usually apparent and can cause diagnostic

difficulty when extensive. Careful inspection of the histology of the necrotic tissue

residues and selective immunostains would permit the diagnosis [19]. The neoplastic

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cells are characteristically epithelial in appearance with large pleomorphic nuclei and

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poorly defined cytoplasm and frequently have a syncytial arrangement. The large

vesicular overlapping nuclei have a fine nuclear membrane and contain one or more

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prominent irregular nucleoli. Mitotic figures are usually numerous.

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The differential diagnosis between metastatic embryonal carcinoma and large cell

undifferentiated carcinoma can be challenging, especially when there is no history of a

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prior primary carcinoma elsewhere [20]. Occurrence of the neoplasm at a young age is an

important clue for the recognition of embryonal carcinoma, and expression of OCT4 and
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SALL4 can confirm the diagnosis [17, 21-25]. Rare morphological variants of embryonal
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carcinoma might be mistaken for seminoma, yolk sac tumor, or the primitive neural

tubules of immature teratomatous elements [18, 26]. The nuclei of embryonal carcinoma
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are larger and more pleomorphic and vesicular than those of yolk sac tumor. Embryonal
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carcinoma is positive for OCT4 and negative for glypican-3, whereas yolk sac tumor is

negative for OCT4 and usually positive for alpha-fetoprotein (AFP) and glypican 3

(Table 2).

5.3. Pure yolk sac tumor

Testicular yolk sac tumor rarely occurs in pure form in adults, but frequently is

seen as a component of a mixed GCT. A highly elevated serum level of AFP is an

important clinical indicator.

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Yolk sac tumor is histologically complex, and various histological patterns have

been described (Fig. 5). These patterns often coexist, but usually one or two are

predominant. The microcystic or reticular pattern is the most common, consisting of a

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network of vacuolated cells resulting in a honeycomb appearance. The macrocystic

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pattern comprises thin-walled cystic spaces of variable size. The solid pattern is

composed of aggregates of polygonal cells with clear or acidophilic cytoplasm, distinct

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cell borders, and small nuclei. The endodermal sinus pattern, the hallmark of yolk sac

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tumor, is found in only 20% of neoplasms. It is characterized by perivascular formations

composed of a stalk of connective tissue containing a thin-walled vessel with a surface

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lined by a layer of cuboidal or low columnar epithelial cells. These structures are known

as SchillerDuval bodies because of their resemblance to the endodermal sinuses of

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Duval present in the rat placenta [27].

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The papillary pattern frequently occurs in association with the endodermal sinus

pattern and consists of numerous fine papillae lined by epithelial cells with a variable
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amount of cytoplasm and enlarged nuclei. The glandular-alveolar pattern is composed of

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gland-like spaces lined by polygonal cells, resembling the human secondary yolk sac at a

gestational age of 67 weeks. This pattern is invariably continuous with microcystic,

endodermal, or papillary patterns. The myxomatous pattern is characterized by abundant

loose myxoid material resembling extraembryonic mesenchyme. It is frequently observed

in testicular yolk sac tumors after chemotherapy.

Polyvesicular vitelline, hepatoid, and glandular or primitive endodermal

(intestinal) patterns are uncommon. The polyvesicular vitelline pattern is more frequent

in ovarian yolk sac tumors, and rarely occurs in pure form. It consists of numerous cysts

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or vesicles lined by a biphasic epithelium, partly flat mesothelial-like cells and partly tall

columnar vacuolated cells [28]. The presence of a constriction in the cyst was

traditionally considered to reflect the process of conversion of the primary yolk sac to the

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secondary yolk sac, but more plausibly, it represents the recapitulation of the allantois

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[29].

The hepatoid pattern is defined as the presence of areas with fetal hepatocytes or

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hepatocellular carcinoma. These cells not only express AFP and hepatocyte paraffin

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antibody-1 (HepPar1), but they also show bile secretion and true hepatic differentiation

ultrastructurally, including bile canaliculus formation. The glandular or primitive

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endodermal (intestinal) pattern is composed entirely of primitive endodermal glands

replicating intestinal or pulmonary glands at an early developmental stage [30]. The

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histological hallmark of these columnar epithelial cells is the presence of subnuclear or

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apical cytoplasmic vacuolation that histologically resemble endometrial glands in early

secretory phase. These cells can express thyroid transcription factor-1. Some intestinal
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yolk sac tumors contain goblet cells that show neuroendocrine differentiation and very
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rarely goblet cell carcinoid formation. Hence, for the purpose of nomenclature, the term

glandular pattern is more appropriate than the others. Some authors refer to the hepatoid

and glandular patterns collectively as the endodermal somatic pattern [29]. Both patterns

occur predominantly in ovarian tumors and occasionally in metastatic testicular germ cell

tumors after chemotherapy.

Another important feature of yolk sac tumor is the presence of diastase-resistant

PAS-positive hyaline globules or bodies. These globules are only occasionally positive

for AFP. They are observed in all yolk sac tumor variants, but occur most frequently in

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the solid and endodermal sinus patterns. They can occur in the cytoplasm of the tumor

cells, vascular spaces, or in the underlying mesenchyme. The analogous PAS-positive,

diastase-resistant material is encountered in mesenchymal connective tissue forming

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bands or patches. The substance usually displays a basement membrane-like

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immunoprofile that includes expression of collagen IV and laminin, but lacks expression

of AFP. The designation parietal differentiation of yolk sac tumor is applied when the

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abundant hyaline basement membrane material becomes a prominent feature [31].

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Occasionally, hematopoiesis can be seen in the capillary spaces. In a minority of yolk sac

tumors, the mesenchymal components can differentiate into smooth muscle, striated
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muscle, or cartilage. Moreover, overgrowth, and even sarcomatous changes of the

mesenchymal component can occur in yolk sac tumors postchemotherapy since cytotoxic
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agents substantially suppress and kill the epithelial cells. Under such unusual
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circumstances, extensive sampling can be required to find the epithelial component. The

detection of isochromosome 12 in the mesenchymal tissue can facilitate the diagnosis of

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postpubertal type yolk sac tumor [32]. The nuclei of yolk sac tumor are often smaller than
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those of seminoma and embryonal carcinoma, and the nuclei are not overlapping, as is

characteristic of embryonal carcinoma.

5.4. Pure choriocarcinoma and other trophoblastic tumors

Although listed in the latest WHO Classification under the category of germ cell

tumors derived from GCNIS, we are not aware of any reported cases of GCNIS

associated with trophoblastic tumors. Nevertheless, their relationship to other malignant

GCTs is confirmed by their occurrence as a component of mixed GCT.

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Choriocarcinoma of the testis is rare and remains uncommon even as a component

of mixed GCT (Fig. 6A) [33]. Unlike other testicular malignant GCTs, choriocarcinoma

most often presents with symptoms related to metastatic disease. Some patients have

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gynecomastia, due to high levels of human chorionic gonadotropin. The majority of

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patients are between 10 and 30 years of age. Neoplasms containing other germ cell

elements in addition to choriocarcinoma, are diagnosed as mixed GCTs but the

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occurrence of a component of choriocarcinoma needs to be mentioned in the diagnosis

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because of its unfavorable prognosis. Choriocarcinomas are positive for hCG, SALL4,

GATA3, glypican-3, alpha-inhibin, and human placental lactogen (Table 2) [34].

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Nonchoriocarcinomatous trophoblastic tumors are exceedingly rare and include

placental site trophoblastic tumor, epithelioid trophoblastic tumor, and cystic

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trophoblastic tumor (Fig. 6B) [35, 36]. They have been reported more commonly as a
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component of mixed GCT or found in metastases after chemotherapy.

5.5. Pure teratoma

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5.5.1. Types of teratoma

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Among GCTs, teratoma arguably displays the greatest diversity in histology,

biological behavior, and terminology between those occurring in the testis and ovary. In

the latest WHO Classification, pure testicular teratoma is divided into two main forms

based in part on the patients age: prepubertal and postpubertal types; however, the

former can occasionally occur at adults, and the latter can occur in prepubertal

individuals who have a disorder of sex development [3]. Postpubertal type teratoma is

more frequent and is associated with GCNIS and chromosome 12p amplification [14, 37,

38]. Postpubertal type teratoma is clinically aggressive, whereas pure prepubertal type

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teratoma has an indolent clinical course. To prevent possible misunderstanding by

clinicians and patients, it might be helpful to add the term malignant in parenthesis to

postpubertal type teratoma and benign to the usual prepubertal type teratoma.

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5.5.2. Postpubertal type teratoma

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Postpubertal type teratoma has pathological features that distinguish it from the

prepubertal type of testicular teratoma as well as ovarian mature cystic teratoma,

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including a predominantly solid appearance on macroscopic examination, more

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disordered arrangement of tissues that are deviated from that of the normal organ, less

presence of thyroid tissue, absence of pituitary gland tissue, more frequent cytological
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atypia, and mitotic figures, and the identification of GCNIS in the residual seminiferous

tubules. There is strong evidence that postpubertal type testicular teratoma arises from a
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transformed germ cell through the intermediary of GCNIS, whereas pure prepubertal type
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testicular teratoma including dermoid cyst, whether prepubertal or postpubertal, as well

as ovarian mature cystic teratoma, arises from a non-transformed germ cell. Cytogenetic
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study of postpubertal type teratoma shows aneuploidy with DNA values in the
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hypotriploid range [14].

About one-third of patients with postpubertal type teratoma present in advanced

stage with metastasis in approximately 20% cases. Postpubertal type teratoma not

uncommonly shows significant cytological atypia and a primitive, mitotically active

cellular stroma that cuffs glands [3]. With rare exceptions, all adult type teratomas are

considered malignant and there is no need to grade or quantify the amount of immature

component (Fig. 7). There is evidence that their so-called fibrous stroma actually

contains a malignant component of the teratoma. It is postulated that somatic type

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malignancy of teratoma is mainly derived from those malignant stromal elements [39].

The finding of concordant chromosome 12p abnormalities between stromal and epithelial

cells from the same tumor suggests that epithelial and fibrous components of teratoma are

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derived from a common progenitor [40-43].

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The primary histological criterion for the distinction between postpubertal and

prepubertal types of teratoma, however, is the presence of GCNIS in the former.

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Postpubertal type teratoma and the fully differentiated cells in residual teratoma post-

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chemotherapy are hypotriploid [3]. Pure postpubertal type teratoma is much less common

than teratoma arising in mixed GCT (see further discussion in Section 5.6 mixed germ
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cell tumor and Section 6.1 Prepubertal type teratoma) [44].

Carcinoid is a low-grade neuroendocrine neoplasm. Most primary testicular

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carcinoids are histologically pure; only about 25% contain other components of teratoma
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(Fig. 7B) [45, 46]. Pure primary testicular carcinoid may represent a burnt-out teratoma

or simply a monodermal teratoma. It may be seen as a pure form (primary carcinoid) or

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in association with teratoma. It can occur in either prepubertal or postpubertal patients,

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though the majority of carcinoids are detected after puberty. The median age is 48 years

(range, 10-83 years [45, 46].

Testicular carcinoids are limited to the insular and trabecular types. They typically

lack associated GCNIS; however, the presence of GCNIS in the surrounding testicular

tissue in occasional cases of pure carcinoid supports the possibility of germ cell origin

[47]. A single study identified the presence of isochromosome 12p in four postpubertal

testicular carcinoids and coexisting teratomas, thus, supporting their germ cell origin

[48]. Primary testicular carcinoid must be distinguished from metastatic carcinoid.

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Limited data indicate that intertubular growth and lymphatic invasion may raise the

possibility of metastasis even when a history of a primary carcinoid elsewhere has not

been established [49].

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On occasion, testicular teratomas have been observed to occur in association with

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somatic-type malignancies other than carcinoid [50]. A secondary malignant component

is encountered only in postpubertal type teratoma and is the result of independent growth

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of an atypical teratomatous embryonic element. The component can be either primitive or

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of adult type. The primitive neoplasms resemble primitive neuroectodermal tumor (Fig.

7C and 7D), nephroblastoma or other blastomatous tumor, embryonal or alveolar

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rhabdomyoosarcoma, or other sarcomas [50-52]. Despite morphologic similarity,

primitive neuroectodermal tumors derived from testicular germ cell tumor lack the classic
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t(11;22) translocation seen in other organ sites [52]. Differentiated rhabdomyomatous

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tumor (Fig. 7F) may occur after chemotherapy for metastatic GCT and should be

distinguished from rhabdomyosarcoma [53]. It should be considered a special form of

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teratoma and has a prognosis similar to that of teratoma.

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A similar pattern of overexpression of 12p and/or isochromosome 12p occurring

in the sarcomatous and the teratomatous component was identified in 21 of 27 testicular

teratomas [39]. This finding supports the origin of the somatic-type malignancy and the

teratoma from the same progenitor cell. Adult type monodermal teratomas other than

carcinoid include adenocarcinoma, or less commonly, squamous cell carcinoma.

5.6. Mixed germ cell tumor

Mixed GCT is a germ cell neoplasm that contains 2 or more malignant GCT

components (Fig. 8A). The most frequent component identified is embryonal carcinoma,

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followed by teratoma, yolk sac tumor, seminoma, and choriocarcinoma. The most

common combinations of components in mixed GCT are embryonal carcinoma with

teratoma, seminoma, and/or yolk sac tumor. Even small amounts of each GCT element

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should be mentioned in the diagnosis because the presence of only a small focus of a

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high-grade component, especially embryonal carcinoma or choriocarcinoma, may greatly

alter the prognosis and require more aggressive treatment. Yolk sac tumor is the most

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often underdiagnosed component. In some examples, the amount of yolk sac tumor is

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very limited. Expression of glypican-3 or AFP is often required to confirm the presence

of a small area of yolk sac tumor. Occasionally, a patient presents with a metastatic
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GCT, and in the testis, there is only macroscopic, histological, or immunohistochemical

evidence of a primary testicular GCT that underwent spontaneous regression [19].

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Teratomatous components that occur in the setting of testicular mixed GCT are
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much more frequent than pure postpubertal type teratoma. The teratomatous elements

show the same allelic losses as those in the other elements, thus, demonstrating the
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common origin of the teratomatous and nonteratomatous components [54]. Metastatic

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teratoma, however, is the most common form of recurrent or metastatic component

following chemotherapy of testicular mixed GCT. Such teratomatous elements frequently

manifest as a persistent or slow-growing mass outside the gonads. The designation

growing teratoma syndrome has been proposed for this situation [55]. Intriguingly, the

primary testicular mixed GCT sometimes does not harbor teratomatous elements, thus

suggesting that teratoma represents terminal differentiation of the mixed GCT.

By definition, the latest WHO Classification of testicular tumors excludes

combined prepubertal type teratoma and yolk sac tumor from the category of mixed GCT

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[3]. Mixed GCT is composed entirely of GCT components; thus, it is distinguished from

gonadoblastoma and mixed germ cellsex cord tumor, both of which also harbor a sex

cord component [56, 57].

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Polyembryoma and diffuse embryoma are rare specialized forms of mixed GCT that

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contain elements of both embryonal carcinoma and yolk sac tumor, but these 2 variants

must be clearly distinguished from each other. Polyembryoma occurs in either the testis

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or the ovary and almost uniformly contains teratomatous elements [14]. It is composed of

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multiple embryoid bodies that resemble a presomite embryo prior to day 18 of gestation

[27]. The embryoid body composed of a central core of embryonal carcinoma and yolk
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sac epithelium projects into a cystic dorsal amnion-like cavity. On the ventral side, yolk

sac tumor epithelium is observed. The embryoid body is embedded in loose myxomatous
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stroma. (Fig. 8B)

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Polyembryoma represents a transitional stage between primitive

(nonteratomatous) and differentiated (teratomatous) GCT and is regarded by some as the

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most immature type of teratoma, recapitulating early embryonic life. An alternative view
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is to consider this tumor as a special form of mixed GCT since it contains both embryonal

carcinoma and yolk sac tumor types of epithelium. The specific pattern is believed to be a

recapitulation of an embryonic structure, the embryonic disc, the center consisting of

embryonal carcinoma-like primitive epithelium overlying thin flattened AFP-positive

yolk sac type epithelium with an amniotic cavity on the dorsal side and the yolk sac

cavity located ventrally. Polyembryoma is only diagnosed as such when such embryoid

bodies become the predominant component in the tumor (>90%). These bodies range

from 0.20.7 mm in greatest dimension. The proliferation of the embryonic disc

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(embryonal carcinoma) or yolk sac component can result in the loss of the classical

embryoid body and should be mentioned in the pathological report as it can have

potential prognostic significance.

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As far as we are aware, diffuse embryoma has only been described in the testis

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[58, 59]. It must be distinguished from polyembryoma and is characterized by an orderly

arrangement of embryonal carcinoma and yolk sac tumor in a necklace-like arrangement

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[14]. The flattened layer usually expresses AFP [58, 59]. Minor components of

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syncytiotrophoblastic cells or teratomatous elements can also be seen.
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6. PREPUBERTAL TYPE GERM CELL NEOPLASMS

The most recent WHO Classification of Tumours of the testis recognizes the
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importance of separating prepubertal types of testicular GCTs from those of the adult
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type [3]. Three closely related forms of prepubertal type tumor are currently recognized:

yolk sac tumor, teratoma, and combined teratoma and yolk sac tumor. It is well
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documented that these neoplasms can present at an age beyond childhood but
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nevertheless maintain distinct characteristics that distinguish them from adult type

testicular GCT [4].

Although the prepubertal types of GCT and spermatocytic tumor both lack an

association with GCNIS and do not have chromosomal abnormalities on the short arm of

chromosome 12, we believe the prepubertal types of GCT are otherwise distinct from

spermatocytic tumor and, thus, the latter is discussed separately.

6.1. Prepubertal type teratoma

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Prepubertal type teratoma is defined as a GCT that usually occurs in the

prepubertal testis and is composed of elements resembling somatic tissues derived from

one or more of the germinal layers, i.e., ectoderm, mesoderm, and endoderm (Fig. 9A)

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[3]. It is histologically similar to mature cystic teratoma of the ovary [60]. Pure

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prepubertal type testicular teratoma has a diploid DNA content and a 46,XY karyotype,

whereas ovarian mature teratoma almost invariably has a normal 46,XX karyotype [61].

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These neoplasms arise directly from a germ cell that has not undergone malignant

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transformation [62]. By definition, the surrounding testis shows no evidence of GCNIS or

chromosome 12p abnormalities. A recent study, however, showed that a small subset of
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prepubertal teratomas can harbor the hallmark genetic alterations (i.e. isochromosome

12p) of adult type teratomas [63]. The significance of this observation has not been
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established.
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Dermoid cyst is a specialized form of prepubertal type teratoma derived entirely

from ectoderm that replicates the structure of skin in an organoid arrangement and
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sometimes also contains neuroectodermal elements (Fig. 9B) [3]. Its occurrence at any
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age can be explained by its slow growth rate. A disadvantage of using the term is that it

fails to connote a neoplastic state to clinicians and patients.

6.2. Epidermoid cyst

For convenience, epidermoid cyst is discussed in this section. Epidermoid cyst is

extremely rare and can occur at any age. In the male genital tract, epidermoid cyst can

involve the testicular parenchyma, the tunica albuginea, or the epididymis (Fig. 9C). It

also occurs in the skin and other sites. On the sectioned surface, it consists of a unilocular

cyst containing lamellar keratin, thus, contrasting with the complex, variegated, solid and

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cystic sectioned surface of most teratomas [14]. On histological examination, it is a

unilocular cyst containing keratin lined by squamous epithelium without skin appendages

or other elements. No GCNIS is identified in the surrounding testicular parenchyma, and

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chromosome 12p abnormalities are nor identified [14, 64]. The neoplastic nature of

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epidermoid cyst or its possible relation to teratoma has not been established [14].

Testicular epidermoid cyst must be distinguished from postpubertal type teratoma. No

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treatment other than simple excision is required, and the clinical outcome is uneventful.

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6.3. Prepubertal type yolk sac tumor

Pure prepubertal yolk sac tumor is the most common testicular GCT in infants
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and children, peaking at the age of 1.5 years (Fig. 9D). An elevated serum level of AFP

beyond what would be expected for the patients age is an important clue for its
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diagnosis. Although histologically identical, there are distinct differences in biological

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behavior between yolk sac tumors that occur in the prepubertal age group and those

occurring after puberty. The absence of GCNIS in the surrounding testicular tubules and
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the absence of chromosome 12p abnormalities indicates that yolk sac tumor in
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prepubertal children arises by a pathway different from those occurring in adults.

Prepubertal type yolk sac tumor has a more favorable prognosis and can be treated less

aggressively than the adult type [65].

6.4. Combined prepubertal type teratoma and yolk sac tumor

Rarely, a minor component of yolk sac tumor occurs in prepubertal type teratoma.

Since these neoplasms have the same excellent prognosis as that of prepubertal type yolk

sac tumor when diagnosed correctly and treated accordingly, they are discussed in this

section. These combined neoplasms are believed to occur as a result of progression of

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prepubertal type teratoma; the teratoma component is diploid, whereas the yolk sac tumor

component is aneuploid [3]. In some examples, the amount of yolk sac tumor is very

limited. Expression of glypican-3 or AFP is often required to confirm the presence of a

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small area of yolk sac tumor. These combined tumors must be distinguished from mixed

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GCTs in postpubertal patients that are associated with GCNIS and amplification of

chromosome 12p.

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7. NEOPLASM WITH GAIN IN CHROMOSOME 9 DUE TO ADDITIONAL

COPIES OF DMRT1 GENE

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7.1. Spermatocytic tumor

In the recently published WHO Classification of Tumours of the Urinary System and
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Male Genital Organs, the term spermatocytic tumor has replaced the older term
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spermatocytic seminoma [3]. Although spermatocytic tumor and prepubertal type GCTs

share a lack of association with GCNIS and lack chromosomal abnormalities on the short
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arm of chromosome 12, we believe that spermatocytic tumor is otherwise distinct from
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the prepubertal types of GCT and, therefore, belongs in a separate category.

Spermatocytic tumor is unique among GCTs because it has only been described in the

testis; it does not occur in the ovary or extragonadal sites. Spermatocytic tumor occurs

predominantly in older patients (average age, 55 years) than classical seminoma although

10% of the former are diagnosed in patients less than 30 years of age [14]. The typical

presentation is a painless scrotal mass. GCNIS is not identified in the residual

seminiferous tubules; however, intratubular growth of spermatocytic tumor is common

and can be confused with the former.

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Histologically, spermatocytic tumor typically is composed of a tripartite

population of cells that vary greatly in nuclear size; the small cells resemble

spermatogonia, whereas the intermediate and large cells resemble primary spermatocytes

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that have entered meiosis 1 (Fig. 10). Uncommonly, spermatocytic tumor has a

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monomorphous appearance, however, and is composed of germ cells with intermediate-

sized nuclei and prominent nucleoli that resemble primary spermatocytes.

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The behavior of pure spermatocytic tumor is typically indolent; it can be cured by

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orchiectomy without additional treatment since metastases are very rare. However, in

those cases that undergo sarcomatous transformation, including rhabdomyosarcoma or

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other high-grade sarcomas, metastases are common and the prognosis is poor [14, 66].

On comparative genetic hybridization, spermatocytic tumor shows a consistent

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gain in chromosome 9 that corresponds to additional copies of the DMRT1 (double sex
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and mab related) gene [67, 68]. Its chromosomal composition is completely different

from that of seminoma and other malignant GCTs. Spermatocytic tumor shows complete
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loss of biparental genomic imprinting and re-establishment of paternal only imprinting

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[3]. Immunohistochemically, these tumors are positive for SALL4, NUT (nuclear protein

in testis), GAGE7, NY-ESO-1; CTAG1B, GAGE1, MAGEA3, MAGEA4, and

MAGEC1; and negative for OCT4, CD30, AFP, PLAP, and hCG. Approximately 50% of

cases are positive for CD117 (c-kit) [16].

The implication is that spermatocytic tumor arises from a germ cell more mature

than a primordial germ cell or spermatogonial stem cell, and the preponderance of

evidence favors origin from the spermatogonium with further maturation of some

transformed cells to primary spermatocytes that have entered mitosis 1 [69, 70].

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8.0 CONCLUSION

Our knowledge of gonadal germ cell tumors has progressed in various ways in the

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last few decades including the description of GCNIS, the discovery of isochromosome

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12p and its importance in the development of invasiveness in germ cell tumors, the

identification of specific stem cell markers for germ cell tumors, the recognition that a

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teratomatous component represents terminal differentiation in a mixed GCT, and the

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description of some novel histological variants of germ cell tumors. Careful morphologic

observations based on detailed clinical information are fundamental to the interpretation

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of perplexing testicular GCTs. Newly available antibodies and molecular diagnostic

studies, when necessary, can contribute to a more specific pathological interpretation in

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challenging cases and increase our knowledge of the pathobiology of these neoplasms.
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Effective integration of classical morphologic observations and emerging novel

molecular studies will likely result in optimal patient care; however, diagnostic pathology
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will continue to play an essential role in this process for the foreseeable future.
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Figure Legends

Fig. 1 Major pathways for the development of testicular germ cell tumors. Rectangle with

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perpendicular corners indicates type of neoplasm. Rectangle with rounded corners

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indicates cell or tissue type, process, or structure. Solid arrow indicates a recognized

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pathway; dashed arrow indicates that the pathway has not been established.

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Fig. 2 Noninvasive germ cell neoplasia of the testis. A, The cells of germ cell neoplasia

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in situ (GCNIS) have enlarged hyperchromatic nuclei with cytoplasmic clearing (original
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magnification x400). B, OCT4 immunostain highlights nuclei of neoplastic cells in

GCNIS(original magnification 400). C, GCNIS with scattered syncytiotrophoblastic cells

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(original magnification x200). D, Intratubular embryonal carcinoma has a glandular

pattern with prominent clefts (original magnification x200).

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Fig. 3 Seminoma. A, Sheets of tumor cells with striking lymphocytic infiltrate along

septa. B, Scattered syncytiotrophobastic cells are seen. C, Prominent granulomatous

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inflammation may obscure neoplastic cells. D, Seminoma cells are relative uniform with

clear cytoplasma and polygonal non-overlapping nuclei. E, Tubular growth pattern. F,

Seminoma may mimic Sertoli cell tumor. G, Seminoma shows discohesive growth and

tumor cells have plasmacytoid apperance. H, Intertubular growth pattern. I, Seminoma

spreads into the rete testis. All images original magnification x200.

Fig. 4 Embryonal carcinoma. A, Tumor cells are highly pleomorphic with syncytial

growth and overlapping nuclei (x200). B, Papillary formations are evident (original

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magnification x200). C, Syncytiotrophoblastic cells are present (original magnification

x200). D, Dual color fluorescence in situ hybridization (FISH) shows isochromosome

12p (arrow), as evidenced by 2 12p signals (green) in close proximity to 1 centromeric 12

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signal (red). The other copies of chromosome 12 (right) show a wild-type pattern, with 1

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green 12p signal and 1 red centromeric signal (original magnification x400).

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Fig. 5 Yoll sac tumor. A and B, Microcystic and macrocystic patterns are intermixed. C,

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Schiller-Duval bodies are noted. D, Glandular pattern is promient. E, Myxoid pattern is

promenent. F, Hepatoid pattern consists of polygonal eosinopilic cells with abundant

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eosinophic cytoplasm. G, Hepatoid cells contain numerous hyaline globues. H,

Sarcomatoid variant of yolk sac tumor is composed of elongated to spindle-shaped cells

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with pleomorphic nuclei. I, Scattered syncytiotroblastic cells are present in tumor with a
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glandular pattern. All images original magnification x200.

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Fig. 6 Trophoblastic tumors. A, Choriocarcinoma consists of sheets of mononuclear

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cytotrophoblasts that are capped by multinucleated syncytiotroblasts (original

magnification x400). B, Cystic trophoblastic tumor is characterized by a cystic space s

lined by squamoid-appearing trophoblasts with prominent cytoplasmic lacunae (original

magnification x200).

Fig. 7 Teratoma and related lesions. A, Postpubertal-type teratoma. Various somatic-type

tissue elements are present (x100). All testicular postpubertal-type teratomas are

potentially malignant regardless of the presence or absence of an immature component.

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B, This teratoma is composed of cartilage only (x200). C, Carcinoid of the testis has a

trabecular growth pattern (x200). D and E, Primitive neuroectodermal tumor (PNET)

with pseudorosette formation (D, original magnification x100; E, original magnification

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x400). F, Differentiated rhabdomyomatous tumor composed of numerous fetal-type

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rhabdomyocytes occurred in a retroperitoneal node dissection after chemotherapy for a

patients previously treated for metastatic mixed germ cell tumor that contained a

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teratomatous component (original magnification x200).

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Fig. 8 Mixed germ cell tumor. A, Tumor is composed of both embryonal carcinoma and
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yolk sac tumor. Scattered syncytiotrophoblasts are also present and should not be

mistaken for choriocarcinoma (original magnification x200). B, Polyembryoma. An

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embryoid body consists of two cavities separated by an embryonic germ disc. The
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centrally located disc is composed of tall epithelium resembling that of embryonal

carcinoma and thin flattened yolk sactype epithelium. The embryoid body is embedded
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in loose myxomatous stroma (original magnification x400).

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Fig. 9 Prepubertal-type germ cell neoplasms. A, Prepubertal-type teratoma. B, Dermoid

cyst. C, Epidermoid cyst. D, Prepubertal-type yolk sac tumor. E and F, Combined

prepubertal-type yolk sac tumor and teratoma. All images original magnification x200.

Fig. 10 Spermatocytic tumor. A, The tumor is composed of a polymorphic tumor

population. Note the presence of three cell types: small cells, intermediate-sized cells,

and large giant cells (original magnification x200). B, Tumor infiltrates between

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seminiferous tubules (original magnification x100). C, Note the presence of pink

eosinophilic materials in large cystic spaces (original magnification x200). D, This

spermatocytic tumor shows solid growth with a uniform appearance of neoplastic germ

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cells with intermediate-sized nuclei. The lack of three distinct cell populations and the

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presence of clear cytoplasm in tumor cells may suggest the diagnosis of seminoma

(original magnification x200). E, Clusters, nests, and single tumor cells are present in a

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tumor with extensive intercellular edema. Tumor cells have a plasmacytoid

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appearance(original magnification x200). F, Another case of spermatocytic tumor with

discohesive growth. Cells have eosinophilic cytoplasm and occasional prominent nucleoli
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(original magnification x200).
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conventional and interphase cytogenetics. Genes Chromosomes Cancer.

1998;23:286-91.

[69] Rajpert-De Meyts E, Jacobsen GK, Bartkova J et al. The immunohistochemical

expression pattern of Chk2, p53, p19INK4d, MAGE-A4 and other selected

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antigens provides new evidence for the premeiotic origin of spermatocytic

seminoma. Histopathology. 2003;42:217-26.

[70] Lim J, Goriely A, Turner GD et al. OCT2, SSX and SAGE1 reveal the phenotypic

T
heterogeneity of spermatocytic seminoma reflecting distinct subpopulations of

P
RI
spermatogonia. J Pathol. 2011;224:473-83.

SC
NU
MA
ED
PT
CE
AC

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PT
RI
SC
NU
MA
ED
PT
CE

Figure 1
AC

38
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PT
RI
SC
NU
MA
ED
PT

Figure 2
CE
AC

39
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PT
RI
SC
NU
MA
ED
PT

Figure 3
CE
AC

40
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PT
RI
SC
NU
MA
ED
PT

Figure 4
CE
AC

41
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PT
RI
SC
NU
MA
ED
PT

Figure 5
CE
AC

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PT
RI
SC
NU
MA
ED
PT
CE
AC

Figure 6

43
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PT
RI
SC
NU
MA
ED
PT
CE
AC

Figure 7

44
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PT
RI
SC
NU
MA
ED
PT
CE
AC

Figure 8

45
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PT
RI
SC
NU
MA
ED
PT

Figure 9
CE
AC

46
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PT
RI
SC
NU
MA
ED
PT
CE
AC

Figure 10

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Table 1 Classification of germ cell tumors of the testis*

________________________________________________________________________
Noninvasive germ cell neoplasia Germ cell neoplasia in situ (GCNIS)
Specific forms of intratubular germ cell

T
neoplasia (specify type)

P
RI
Invasive postpubertal type germ cell Seminoma

SC
tumors Embryonal carcinoma
Yolk sac tumor
Teratoma

NU
Teratoma with somatic type malignancy
MA Choriocarcinoma
Nonchoriocarcinomatous trophoblastic tumor
Placental site trophoblastic tumor
Epithelioid trophoblastic tumor
ED

Cystic trophoblastic tumor

Mixed germ cell tumor
PT
CE

Invasive prepubertal type germ cell Prepubertal type yolk sac tumor
AC

tumors Prepubertal type teratoma

Dermoid cyst
Epidermoid cyst
Combined prepubertal type teratoma and yolk
sac tumor

Spermatocytic tumor

Modified from the 2016 WHO Classification of Tumours of the Testis (3)

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Table 2 Useful immunohistochemical stains in the differential diagnosis of

testicular germ cell tumors

T
E C A H S Mel
AE SA PL OC GA 2- CD CD GP SO SO Inih Calre FO
M K F C F an-
1/3 LL4 AP T4 TA3 4 117 30 C3 X2 X17 ibin tinin XL2

P
A 7 P G -1 A
0

RI
GCNIS - - - + + + - + + - - - - + - - - U U U
/+

SC
Seminom - V - + + + - + + - - - - + - - - - - -
a /+

Embryon - + - + + + - - - + - - + - - - - - - -

NU
al /+
carcinom
a

Yolk sac - + - + + - + - -/+

MA- + + - + - - - - - -
tumor

Choriocar + + + + + - + V - - + - - - + - - - - -
cinoma
ED

Teratoma + + + + V -/+ -/+ - - -/+ - - V V - - - - - -

/+

Spermato - - U + - - - - +* - U - U U - - - U U -
PT

cytic
tumor

Sertoli -/+ V - - - - - - - - - - - U - + V + + -
CE

cell tumor

Leydig - - - - - - - - - - - - - U - + + + -/+ +
cell tumor
AC

JGCT -/+ - - - - -/+ U - - - - - - U - + + + + -

/+

AGCT - - - - - -/+ U - - - - - - U - + + + + -/+

/+

GCNIS, germ cell neoplasia in situ; JGCT, juvenile granulosa cell tumor; AGCT, adult
granulosa cell tumor; EMA, epithelial membrane antigen; PLAP, placental-like alkaline
phosphatase; D2-40, podoplanin; GPC3, glypican 3; HCG, human chorionic
gonadotrophin; SF-1, steroidogenic factor 1; FOXL2, forkhead box L2.

-/+ usually negative; V, variable; U, unknown; *approximately 50% of spermatocytic

tumors are positive for CD117 (c-kit).

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