Seminoma Pathology

Seminoma is the most common pure germ cell tumor (GCT) of the testis, accounting for up to 50% of cases. ^[1] Among mixed GCTs, seminoma is also commonly present in combination with teratoma, yolk sac tumor, and/or embryonal carcinoma. Such tumors are still classified as mixed GCT, even if the seminoma is the main component. Overall, 60% of germ cell neoplasms have seminoma in their composition, but pure seminomas are genetically different from those that present as a component of a mixed tumor. ^[2]

Testicular GCT is the most prevalent solid malignancy affecting young male adults (most commonly from age 30 to 49 years). However, reports of children (8 y) and elderly (73 y) also appear in the literature. ^{[3, 4]}

For unclear reasons, the overall incidence of seminoma has increased progressively in the 21st century in several different countries. ^[5] There are well described racial and geographic differences in the incidence of GCTs. Overall, the incidence of seminoma is 6- to 10-fold higher in White than Black persons, which is observed in all age groups. ^[6] Interestingly, high economic status represents an important association with the development of GCTs overall and seminoma specifically. ^{[7, 8]}

Well-defined associations of seminoma are: cryptorchidism, familial history or a previous testicular GCT, certain intersex syndromes, and oligospermic infertility. ^{[9, 10]} Cryptorchidism represents the most strong risk factor for seminoma, with an increased risk not only in the affected testis, but also in the contralateral one. Recently suggested associations include high estrogen levels, previous radiotherapy or chemotherapy, exposure to organochlorine, ^[11] abusive use of marijuana, ^[12] testicular microlithiasis, and nutrition status during infancy, but these reports lack confirmation. ^[13]

Genetic changes have also been studied in the past few decades, with documentation of aneuploid DNA content in seminomas and intratubular germ cell neoplasia of the unclassified type (IGCNU), the precursor lesion. ^[14] A consistent chromosomal change is present in about 80% of seminomas, which consists of a gain in isochromosome 12p [i(12p)] (see Molecular/Genetics, below).

Most seminomas are intratesticular in location. However, GCTs, including seminomas, can occur in extragonadal sites along the midline of the body, following the embryologic migration route of its precursor cells -- the primordial germ cells. ^[15] Common extragonadal sites include the mediastinum ^{[16, 17]} and central nervous system (CNS) (germinomas). Differences in behavior and clinical outcome suggest that mediastinal seminomas are biologically different from gonadal seminomas and can develop de novo without a testicular primary focus. In contrast, tumors arising in the retroperitoneum are virtually always associated with premalignant lesions in one of the testes and behave clinically similar to metastatic testicular seminomas. ^{[18, 19]}

The mean age at presentation of patients with seminoma is 40 years, which is 5-10 years older than in patients with nonseminomatous testicular tumors. Up to 90% of affected patients present with symptoms and swelling, or other palpable abnormalities are the most common. Pain, however, is an uncommon symptom.

Elevated serum human chorionic gonadotropin (hCG) levels can occur in seminomas and correlate with syncytiotrophoblastic giant cells seen histologically. Serum hCG is mildly to moderately elevated in about 10% of patients with clinical stage I seminoma and 25% of patients with metastasis. Even so, gynecomastia is a rare event. Alpha fetoprotein (AFP) is not produced by seminoma cells, and its serum detection usually indicates a nonseminomatous component. Minimal AFP elevations, however, may be due to hepatopathy, including metastatic seminoma to the liver. ^[20]

The testis is usually enlarged, and the tumor nodule may distort the external appearance of the organ (see the image below). The cut surface is cream-colored to gray-tan to pink. The usual presentation is a lobulated, single to multinodular mass, confined to the testis but with a tendency to bulge into the surrounding parenchyma. Local invasion, if present, is most common in the mediastinum. Extensive hemorrhage or necrosis are unusual but may be present in large tumors. Tumors with predominant interstitial growth pattern or extensive fibrosis may be unapparent or have a scarring appearance.

Seminoma pathology. Pure seminoma. The tumor is white tan and slightly lobulated with foci of hemorrhage. It replaces most of the testicular parenchyma but is organ confined.

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Seminomas often have a diffuse, sheetlike pattern or confluent multinodular pattern. Less frequently, a tubular morphology can be seen (see the images below), posing a differential diagnosis with Sertoli cell tumor.

Seminoma pathology. Low magnification of seminoma replacing testicular parenchyma. Note the few isolated residual tubules.

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Seminoma pathology. Low magnification showing sharp demarcation between seminoma and normal testis. This is distinct from case above, in which the seminoma cells permeated between seminiferous tubules.

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Rare histologic variants of seminomas are cribriform, alveolar, and microcystic. In these cases, seminoma-type nuclei are characteristic and should help with the differential diagnosis. The periphery often shows an interstitial or cordlike pattern, permeating benign seminiferous tubules (intertubular growth pattern). Edema may yield a microcystic or cribriform architecture, mimicking yolk sac tumor.

Dense fibrous septae are common and areas of scarring may occur to a greater or lesser extent. Extensive fibrosis may disguise the diagnosis and complete regression is not infrequent ("burn-out"), as seen in the following image. ^{[21, 22]} Granulomatous reaction is also associated with this phenomenon and, when abundant, it can be misperceived as granulomatous orchitis.

Seminoma pathology. Dense hyalinized tissue is occasionally the only finding in a "burn out" tumor either in the primary testicular site or in the metastatic site, usually the retroperitoneum. Rarely, once can see "ghosts" of malignant germ cells (not shown).

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Seminoma cells have clear to eosinophilic cytoplasm with well defined borders, central nuclei, and 1 or 2 large nucleoli (see the image below). Abundant cytoplasm cause the nuclei to be well spaced although closely apposed. Variations in cytology may output a plasmacytoid appearance. Seminomas contain glycogen and are therefore positive for periodic acid-Schiff (PAS) staining, as opposed to spermatocytic seminoma.

Seminoma pathology. At this power, the nuclear details of seminoma cells can be appreciated. Note the similar cell size; the nuclear anaplasia, which is less than that of embryonal carcinoma; the centrally placed nucleoli; and the pink, moderately abundant cytoplasm.

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Sprinkling lymphocytes are virtually always associated with seminoma and represent a helpful diagnostic feature (see the following image). These are more prominent in and around fibrous trabeculae but are also admixed with seminoma cells. Germinal center formation has a cytotoxic effect, hence, they are usually composed of CD3+ T-cells. ^[23] Larger scattered syncytiotrophoblastic giant cells are seen in up to 20% of seminomas.

Seminoma pathology. Solid growth pattern of seminoma cells with lymphocytic infiltrates.

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Placental alkaline phosphatase (PLAP) is positive in a membranous pattern in up to 98% of seminomas, but this marker is also frequently expressed in other testicular GCTs, except for spermatocytic tumor. In contrast, CD117/c-KIT is specific for seminoma and positive in the majority of cases (see the image below), but it rarely can also be positive in embryonal carcinomas. ^{[24, 25]}

Seminoma pathology. CD117 (KIT) stain showing strong membranous positivity in seminoma cells.

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Other specific markers include podoplanin, SOX17, SALL4, OCT3/4 (see the following two images) and D2-40 that stain seminomas and embryonal carcinomas with 100% sensitivity. ^{[26, 27]} CD30 is negative in seminoma while positive in embryonal carcinomas. ^{[28, 29]} Novel markers not yet widely used in clinical practice are activator protein-2gamma (AP-2gamma), which shows strong sensitivity for seminoma and IGCNU, and transcription factors NANOG and SOX2, which show staining and specificity similarities with OCT3/4. Melanoma-associated gene C2 (MAGEC2) is a newly identified marker that is also useful in distinguishing between seminomas and embryonal carcinomas. MAGEC2 is present in 94% of seminomas, but no expression has been found in embryonal carcinomas, making it another tool that can be used in the diagnosis of testicular germ cell tumors. ^[30]

Seminoma pathology. OCT3/4 immunohistochemical study at high magnification showing strong nuclear reactivity.

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Seminoma pathology. Immunohistochemical stain for OCT3/4 showing seminoma cells with pagetoid spread to the rete testis. This tumor was a large seminoma (not shown) with extensive intratubular germ cell neoplasia, unclassified.

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In a study of 67 cases of pure seminoma of the testis, Renshaw and Gould found evidence that OCT4 staining of lymphohistiocytic inflammation (LHI) near vessels in cases of pure seminoma improves detection of lymphatic/vascular invasion (LVI). They found that prominent LHI may obscure LVI in the spermatic cord of patients with pure seminoma and that OCT4 staining of such foci may increase the rate of LVI detection by 45%. ^[31]

Scattered dot-like keratin can be expressed in up to 36% of seminomas (whereas they are diffuse and strong in embryonal carcinomas). AE1/AE3, CAM5.2, and CK7 can be focally positive in seminomas, whereas CK20 and high molecular weight keratin (HMWK) are negative. Epithelial membrane antigen (EMA) has an expression rate of around 10% in seminomas and can be used along with hCG in the differential diagnosis with choriocarcinoma, in which it is positive in about 50% of cases. ^{[32, 33]}

Testicular GCTs are the neoplastic counterpart of primordial germ cells/gonocytes. Primordial germ cells (PGCs) are derived from the epiblast and migrate to the genital ridge as early as the 6th week of the embryonic development. PGCs are positive for OCT-4, which is thought to regulate pluripotency. Once in the genital ridge, PGCs are called gonocytes and will differentiate into oocytes (ovary) or prespermatogonia (testis). This migration is accompanied by proliferation and is controlled by the KIT stem-cell signaling pathway. ^{[34, 35]}

During normal maturation of germ cells, OCT-4, PLAP, and c-KIT immunoexpression disappear. Nevertheless, the corresponding genes are later reprogrammed during oncogenesis, and these proteins are detected in germ cell neoplasms, including seminomas. ^[9]

KIT gene mutations are implicated in extragonadal survival of PGCs. In view of its presence in bilateral tumors, the mutation must take place in PGCs before their arrival in the gonadal ridges. ^[36]

The initial event in the origin of seminoma is the malignant transformation of an intratubular germ cell. This process is analogous to intraepithelial or carcinoma in situ in other organs, but as gonocytes are not epithelial cells, the accurate terminology is intratubular germ cell neoplasia, unclassified (ITGCNU). Seminomas show relative overrepresentation of 12p chromosome sequences, but no consistent gain of 12p is detected in ITGCNU. These data indicate that overrepresentation of 12p is required for progression from preinvasive to invasive behavior. Candidate genes on 12p include KRAS,CCND2, and NANOG. ^[37]

The default pathway of other testicular GCT is hypothesized to be towards the development of seminoma. A nonseminomatous tumor is thought to require activation of pluripotency (reprogramming) of either a seminoma cell or an ITGCNU cell. DNA indexes are hypertriploid in seminomas, in contrast to nonseminomatous tumors (hypotriploid) possibly due to net loss of chromosomal material during cancer progression. ^[15] Furthermore, seminomas have demethylated DNA compared to nonseminomatous tumors. ^[38]

SOX2 is repressed in seminomas. Kushwaha et al found evidence that SOX2 repression in seminoma-derived TCam-2 cells is mediated by the Polycomb repressive complex (PcG) and the repressive H3K27me3 chromatin mark that are enriched at its promoter. ^[39]

All GCTs are aneuploid most likely because of early establishment of polyploidy. Polyploidy causes genomic instability with consequent genetic heterogeneity. This will lead to the phenotypic changes that drive selective survival of the best adapted clone, along with the morphologic variability of such tumors, as previously discussed.

An abstract presented at the 2015 Genitourinary Cancers Symposium theorized that serum levels of microRNA-371a-3p (miR-371) can be a useful biomarker in testicular GCTs. ^{[40, 41, 42]}

The main staging system for testicular cancers, including seminoma, is the American Joint Committee on Cancer (AJCC) Tumor-Node-Metastasis (TNM) staging protocol of testicular neoplasms. The most important features are vascular invasion, extension of the tumor through the tunica albuginea, which separates pT1 to pT2, invasion of the spermatic cord (pT3), and invasion of the scrotum (pT4). However, different from other organs, the TNM system of testicular tumors adds the particularity of considering serum tumor markers (eg, lactate dehydrogenase [LDH], hCG, AFP). ^[43]

The most important prognostic factor in seminomas is pathologic TNM (pTNM) stage. Volume of primary tumor and rete testis invasion are believed to be of prognostic relevance as well. Modern therapy is eminently stage-dependent and has rendered testicular seminomas highly curable, with overall survival rates exceeding 95%. ^{[44, 45]} Seminomas metastasize most commonly to retroperitoneal nodes and then to lungs. Bone metastases, although infrequent, are more common in seminomas than in nonseminomatous tumors.

A subset of more aggressive-looking tumors, the so-called "anaplastic seminomas," are a group of neoplasms that demonstrate increased nuclear pleomorphism, prominent nucleoli, and 3 or more mitoses per high-power field (HPF) (also called "seminoma with high mitotic rate"). These terms are derived from early studies performed at the Walter Reed Medical Center in Washington, DC. ^[46] In 2002, Tickoo et al described a subset of seminomas with atypical features that tended to present at a higher tumor stage and were less responsive to therapy. ^[47] It is not clear, however, that aggressive treatment is indicated, and it is still a matter of debate if "anaplastic seminomas" are more aggressive stage-wise when compared with the classical counterpart.

Vascular invasion is a predictor of recurrence in nonseminomatous primary neoplasia, but its true significance is not well established in seminomas. ^[48] Vascular invasion demands cautious evaluation; once seminoma cells are easily detached during tissue processing, they tend to get artifactually placed in vascular spaces ("knife artifact"). True vascular invasion should display seminoma cells admixed with blood cells and/or adherent to the vessel wall. In selected cases, immunohistochemistry for endothelial cells (CD31, CD34) may be helpful.