Genome Wide DNA Methylation Profiles Provide Clues to the Origin and Pathogenesis of Germ Cell Tumors

The cell of origin of the five subtypes (I-V) of germ cell tumors (GCTs) are assumed to be germ cells from different maturation stages. This is (potentially) reflected in their methylation status as fetal maturing primordial germ cells are globally demethylated during migration from the yolk sac to the gonad. Imprinted regions are erased in the gonad and later become uniparentally imprinted according to fetal sex. Here, 91 GCTs (type I-IV) and four cell lines were profiled (Illumina’s HumanMethylation450BeadChip). Data was pre-processed controlling for cross hybridization, SNPs, detection rate, probe-type bias and batch effects. The annotation was extended, covering snRNAs/microRNAs, repeat elements and imprinted regions. A Hidden Markov Model-based genome segmentation was devised to identify differentially methylated genomic regions. Methylation profiles allowed for separation of clusters of non-seminomas (type II), seminomas/dysgerminomas (type II), spermatocytic seminomas (type III) and teratomas/dermoid cysts (type I/IV). The seminomas, dysgerminomas and spermatocytic seminomas were globally hypomethylated, in line with previous reports and their demethylated precursor. Differential methylation and imprinting status between subtypes reflected their presumed cell of origin. Ovarian type I teratomas and dermoid cysts showed (partial) sex specific uniparental maternal imprinting. The spermatocytic seminomas showed uniparental paternal imprinting while testicular teratomas exhibited partial imprinting erasure. Somatic imprinting in type II GCTs might indicate a cell of origin after global demethylation but before imprinting erasure. This is earlier than previously described, but agrees with the totipotent/embryonic stem cell like potential of type II GCTs and their rare extra-gonadal localization. The results support the common origin of the type I teratomas and show strong similarity between ovarian type I teratomas and dermoid cysts. In conclusion, we identified specific and global methylation differences between GCT subtypes, providing insight into their developmental timing and underlying developmental biology. Data and extended annotation are deposited at GEO (GSE58538 and GPL18809).     

In vitro transformation of mouse testis cells by oncogene transfection

Germ cell tumors (GCTs) are unique in that they exhibit diverse biological characteristics and pathological features. Although several in vivo GCT models are available, studies on GCTs are hampered because in vivo development of GCTs is time consuming and prevents a detailed molecular analysis of the transformation process. Here we developed a novel strategy to transform mouse testis cells in vitro. Lentivirus-mediated transfection of dominant negative Trp53, Myc, and activated Hras1 into a CD9-expressing testis cells caused tumorigenic conversion in vitro. Although these cells resembled embryonic stem (ES) cells, they were aneuploid and lacked Nanog expression, which is involved in the maintenance of the undifferentiated state in ES cells. Euploid ES-like cells were produced by transfecting the Yamanaka factors (Pou5f1, Myc, Klf4, and Sox2) into the same cell population. Although these cells expressed Nanog, they were distinct from ES cells in that they expressed CD44, a cancer stem cell antigen. Both treatments induced similar changes in the DNA methylation patterns in differentially methylated regions of imprinted genes. Moreover, despite the differences in their phenotype and karyotype, both cell types similarly produced mixed GCTs on transplantation, which were composed of teratomas, seminomas, and embryonal carcinomas. Thus, in vitro testis cell transformation facilitates an analysis of the GCT formation process, and our results also suggest the close similarity between GCT formation and reprogramming.     

The plasticity of germ cell cancers and its dependence on the cellular microenvironment

So far, the understanding of germ cell cancer (GCC) pathogenesis is based on a model, where seminomas and non‐seminomas represent distinct entities although originating from a common precursor termed germ cell neoplasia in situ (GCNIS). Embryonal carcinomas (ECs), the stem cell population of the non‐seminomas, is pluri‐ to totipotent and able to differentiate into cells of all three germ layers, giving rise to teratomas or tumours mimicking extraembryonic tissues (yolk sac tumours, choriocarcinomas). With regard to gene expression, (epi)genetics and histology, seminomas are highly similar to GCNIS and primordial germ cells, but limited in development. It remains elusive, whether this block in differentiation is controlled by cell intrinsic mechanisms or by signals from the surrounding microenvironment. Here, we reviewed the recent literature emphasizing the plasticity of GCCs, especially of seminomas. We propose that this plasticity is controlled by the microenvironment, allowing seminomas to transit into an EC or mixed non‐seminoma and vice versa. We discuss several mechanisms and routes of reprogramming that might be responsible for this change in the cell fate. We finally integrate this plasticity into a new model of GCC pathogenesis, allowing for an alternative view on the dynamics of GCC development and progression.     

Detection of chromosome aberrations in paraffin sections of seven gonadal yolk sac tumors of childhood

Yolk sac tumors are the most frequent kind of malignant pediatric germ cell tumor and may have a fundamentally different pathogenesis than adult germ cell tumors. Since few cytogenetic studies have been performed so far, in situ hybridization was applied to interphase cell nuclei of seven gonadal yolk sac tumors of childhood in routine paraffin-embedded tissue sections. The panel of chromosome-specific DNA probes was selected on the basis of their relevance in adult germ cell tumors and consisted of five DNA probes specific for the (peri)centromeric regions of chromosomes 1, 8, 12, 17 and/or X and/ or one DNA probe specific for the subtelomeric region of chromosome 1 (p36.3). As in adult germ cell tumors, all pediatric gonadal yolk sac tumors had an increased incidence of numerical chromosome aberrations. All tumors showed an overrepresentation of at least three chromosomes. Gains of chromosome 12, which is highly specific in adult germ cell tumors, were diagnosed in six pediatric gonadal yolk sac tumors. The DNA indices determined in the paraffin-embedded tumor material correlated well with the in situ hybridization findings. A chromosome was either over- or underrepresented, compared with the corresponding DNA indices, in only a few cases. The short arm of chromosome 1 in adult germ cell tumors is often involved in structural aberrations. In pediatric germ cell tumors, the short arm of chromosome 1 is also a nonrandom site of structural aberrations. Moreover, the presence of a deletion at 1p36.3 in four out of five tumors suggests that the loss of gene(s) in this region is an important event in the pathogenesis of gonadal yolk sac tumors of childhood.     

α-Fetoprotein-Producing Non-Germ Cell Tumors of the Urological System

Elevated serum levels of α-fetoprotein (AFP), a fetal serum protein, occur mainly in the development of hepatocellular carcinoma (HCC) or germ cell tumors, mainly yolk sac tumor. Rarely, other tumors of the urological system produce AFP. This article reviews the AFP-producing non-germ cell tumors of the urological tract reported to date. These include different types of tumors of the adrenal glands, kidney, ureter, urinary bladder, and testis. It is important for pathologists, urologists, and oncologists to be aware of such cases as the diagnosis affects the management plan for the patient.     

Risk of malignant childhood germ cell tumors in relation to demographic,gestational, and perinatal characteristics

BackgroundChildhood germ cell tumors (GCTs) are a rare assortment of neoplasms, with mostly unknown etiology, that are believed to originate very early in life. Few studies have examined risk factors by histologic subtype, despite evidence of different risk profiles.Materials and methodsIn this population-based case-control study, 451 childhood malignant GCT cases ages 0–5 years were identified from the California Cancer Registry. Differentiating between common histologic subtypes, we identified 181 yolk sac tumors, 216 teratomas, and 54 rarer subtypes. Cases were linked to their birth certificates and 271,381 controls, frequency matched by birth year, were randomly selected from California birthrolls to investigate the contributions of demographic, gestational, and pregnancy factors using unconditional logistic regression analysis.ResultsCompared to non-Hispanic whites, Asian/Pacific Islander children were at an increased risk for developing GCTs (odds ratio [OR] = 1.94; 95% confidence interval [CI] = 1.47, 2.56). Among pregnancy complications and procedures, yolk sac tumors were positively associated with the presence of fetopelvic disproportion (OR = 2.97; 95% CI = 1.55, 5.68), while teratomas were strongly associated with polyhydramnios or oligohydramnios (OR = 14.76; 95% CI = 7.21, 30.19) and the presence of an ear, face, or neck anomaly at birth (OR = 93.70; 95% CI = 42.14, 208.82).ConclusionsMalignant yolk sac tumors and malignant teratomas exhibited distinct demographic and gestational characteristics; additionally, complications in pregnancy and labor may be brought on by specific histologic subtypes.     

A mini-review of familial ovarian germ cell tumors: An additional manifestation of the familial testicular germ cell tumor syndrome

Introduction While testicular germ cell tumors (TGCTs) are the most common malignancy in young men, germ cell tumors in women are uncommon. Familial clustering, epidemiologic evidence of increased risk with family or personal history of TGCT, and associations with genitourinary tract anomalies suggest an underlying genetic predisposition to TGCT, but traditional linkage studies have yet to identify a highly penetrant TGCT cancer susceptibility gene. In this paper, we investigate the familial occurrence of testicular and ovarian germ cell tumors. Methods We report a family in which a TGCT and an ovarian germ cell tumor (OGCT) occurred in two siblings, summarize the existing literature on familial occurrences of OGCT, either alone or in combination with extragonadal or TGCTs, and compare the incidence of familial and sporadic testicular and ovarian GCTs. Sporadic GCT data were obtained from the US Surveillance Epidemiology and End Results (SEER) registry. Results We identified 16 reports of OGCT occurring in conjunction with either ovarian, testicular or extragonadal GCT. In these familial cases, the mean age at onset of female dysgerminoma was younger than that noted in the general population (age 17 vs. age 24, p = 0.01). In SEER, the incidence of TGCT was 15 times higher than that of OGCT. Histologic distributions in males and females showed distinctly different patterns. Discussion Although the incidence of OGCTs in the general population is quite low, its occurrence in multiple members of the same family and in families with TGCT suggests that a gene conferring susceptibility to GCTs may exist in some families.     

Molecular biomarkers as potential targets for therapeutic strategies in human testicular germ cell tumors: An overview

Testicular germ cell tumors (TGCTs), the most common malignancy in males between 15 and 34 years of age and the most frequent cause of death from solid tumors in this age group. TGCTs can be subdivided into seminoma and non‐seminoma germ cell tumors (NSGCTs), including embryonal cell carcinoma, choriocarcinoma, yolk sac tumor, and teratoma. Seminomas and NSGCTs do not only present distinctive clinical features, but they also show significant differences as far as therapy and prognosis are concerned. Seminomas are highly sensitive to both radiation and chemotherapy, with a good prognosis, non‐seminomas are sensitive to platinum‐based combination chemotherapy and are less susceptible to radiation, with the exception of teratomas. The different therapeutic outcome might be explained by inherent properties of the cells from which testicular neoplasia originate. The unique treatment sensitivity of TGCTs is unexplained so far, but it is likely to be related to intrinsic molecular characteristics of the PGCs/gonocytes, from which these tumors originate. Many discovered bio‐markers including OCT3/4, SOX2, SOX17, HMGA1, HMGA2, PATZ1, GPR30, Aurora B, estrogen receptor β, and others have given further advantages to discriminate between histological subgroups. In addition, therapeutic approaches for the treatment of TGCTs have been proposed: humanized antibodies against receptors/surface molecules on cancer cells, inhibitors of serine–threonine, and tyrosine kinases, and others. The mini‐review will be an overview on the molecular alterations identified in TGCTs and on novel targeted antineoplastic strategies that might help to treat chemotherapy resistant TGCTs. J. Cell. Physiol. 228: 1641–1646, 2013. © 2013 Wiley Periodicals, Inc.     

The Hypoxia-Inducible Epigenetic Regulators Jmjd1a and G9a Provide a Mechanistic Link between Angiogenesis and Tumor Growth

Hypoxia promotes stem cell maintenance and tumor progression, but it remains unclear how it regulates long-term adaptation toward these processes. We reveal a striking downregulation of the hypoxia-inducible histone H3 lysine 9 (H3K9) demethylase JMJD1A as a hallmark of clinical human germ cell-derived tumors, such as seminomas, yolk sac tumors, and embryonal carcinomas. Jmjd1a was not essential for stem cell self-renewal but played a crucial role as a tumor suppressor in opposition to the hypoxia-regulated oncogenic H3K9 methyltransferase G9a. Importantly, loss of Jmjd1a resulted in increased tumor growth, whereas loss of G9a produced smaller tumors. Pharmacological inhibition of G9a also resulted in attenuation of tumor growth, offering a novel therapeutic strategy for germ cell-derived tumors. Finally, Jmjd1a and G9a drive mutually opposing expression of the antiangiogenic factor genes Robo4, Igfbp4, Notch4, and Tfpi accompanied by changes in H3K9 methylation status. Thus, we demonstrate a novel mechanistic link whereby hypoxia-regulated epigenetic changes are instrumental for the control of tumor growth through coordinated dysregulation of antiangiogenic gene expression.     

Bilateral germ cell testicular tumors

PURPOSE: To study the clinical characteristics of bilateral testicular tumors in the cisplatin era. PATIENTS AND METHODS: Between November 1988 and November 1998 2386 testicular cancer patients were treated in our Department and 72 bilateral germ cell testicular cancer patients were retrospectively explored (3%). The incidence, the clinical and histological characteristics and, in the case of asynchronous tumor, the interval between the two tumors were analyzed. RESULTS: During the 10 years 19 synchronous (26.4%) and 53 asynchronous bilateral germ cell testicular cancers (73.6%) were treated. The incidence of bilateral synchronous seminoma was 68.4%. Among the asynchronous tumors 9 concordant seminomas and 9 concordant nonseminomas were detected. In the first, second and third 5-year follow-up period 39.6, 30.2, and 28.2% of asynchronous tumors were diagnosed. The incidence of seminoma after the first castration in the 5, 10 and 15 years was 19, 37.5, and 60%, respectively. The overall survival rates of synchronous and asynchronous testicular cancer were 84 and 93%. In cases of asynchronous tumor the prevalence of stage I cancer was significantly greater in a regularly controlled population (p=0.014) than in the not regularly followed population, but the survival rate was good in both groups. Nonseminoma showed up earlier as first and second tumor than seminoma (p=0.05, p=0.045). The interval between the two asynchronous tumors was shorter in the case of nonseminoma than in the case of seminoma (p=0.002). CONCLUSION: The prognosis of bilateral germ cell testicular cancer is good because of the high incidence rate of seminoma and the effective treatment. With regular follow-up the early diagnosis of second testicular tumors is probable. The interval between the tumors depends on the patients' age and the histology of the second tumor, in the case of seminoma it is longer. The effect of the previous treatment on the incidence of seminoma and the interval between the two asynchronous tumors requires further investigations.     

Aurora B expression in post-puberal testicular germ cell tumours

Aurora/Ipl1-related kinases are a conserved family of proteins that are essential for the regulation of chromosome segregation and cytokinesis during mitosis. Aberrant expression and activity of these kinases occur in a wide range of human tumours and have been implicated in mechanisms leading to mitotic spindle aberrations, aneuploidy, and genomic instability. Previous studies of our group have shown that Aurora B expression is restricted to specific germinal cells. In this study, we have evaluated by immunohistochemical analysis Aurora B expression in post-puberal testicular germ cell tumours (22 seminomas, 2 teratomas, 15 embryonal carcinomas, 5 mixed germinal tumours with a prominent yolk sac tumour component and 1 choriocarcinoma). The Aurora B protein expression was detected in all intratubular germ cell tumours, seminomas and embryonal carcinomas analysed but not in teratomas and yolk sac carcinomas. The immunohistochemical data were further confirmed by Western blot analysis. In addition, the kinase Aurora B was vigorously expressed in GC-1 cells line derived from murine spermatogonia. The block of Aurora B function induced by a pharmacological inhibitor significantly reduced the growth of GC-1 cells suggesting that Aurora B is a potential therapeutic target. J. Cell. Physiol. 221: 435–439, 2009. © 2009 Wiley-Liss, Inc.     

Differentiation of human testicular embryonal carcinoma and teratocarcinoma grown in nude mice and soft-agar cultures

The differentiation pattern of two human germ cell tumors, grown in nude mice and in vitro is described. Tumor A was an embryonal carcinoma (EC) of borderline histology with characteristics of yolk sac tumor and of seminoma; tumor B was a teratocarcinoma with yolk sac elements and syncytiotrophoblastic giant cells. The morphology of an EC as well as cytogenetic characteristics were maintained during 20 passages in nude mice from tumor A and over 11 passages from tumor B. Tumor A did not grow in vitro. Cell suspensions prepared from xenografted tumor B grew into cystic embryoid bodies in semi-solid tissue culture medium. These embryoid bodies showed cuboidal and flattened cells with microvilli, junctional complexes, peripheral microfilaments, and annulated lamellae, reminiscent of the 'inner cell mass' of a blastula and of endoderm, respectively. When such colonies were transplanted into nude mice, however, only tumors with the morphology found in the transplants appeared.     

Survival after a diagnosis of testicular germ cell cancers in Germany and the United States, 2002–2006: A high resolution study by histology and age

Introduction: The aim of this study was to provide detailed age-specific (5-year age groups) and histology-specific (histologic subtypes of seminoma and nonseminoma) relative survival estimates of testicular germ cell cancer patients in Germany and the United States (U.S.) for the years 2002–2006 and to compare these estimates between countries. Methods: We pooled data from 11 cancer registries of Germany and used data from the U.S. (SEER-13 database) including 11,508 and 10,774 newly diagnosed cases (1997–2006) in Germany and the U.S., respectively. We estimated 5-year relative survival (5-year-RS) by histology and age based on period analysis. Results: 5-year-RS for testicular germ cell tumors was 96.7% and 96.3% in Germany and the U.S., respectively. 5-Year-RS for spermatocytic seminoma was close to 100% in both countries. 5-Year-RS for nonseminoma was lower than for classical seminoma in Germany (93.3% versus 97.6%) and the U.S. (91.0% versus 98.2%). Among nonseminomas, choriocarcinomas provided the lowest 5-year-RS in both countries (Germany 80.1%, U.S. 79.6%). Age-specific 5-year-RS for seminoma showed only little variation by age. 5-Year-RS for nonseminomas tended to be lower at higher ages, especially for malignant teratoma. Discussion: This is the first study that provides up-to-date survival estimates for testicular cancer by histology and age in Germany and the U.S. Survival after a diagnosis of testicular cancer is very comparable between Germany and the U.S. 5-Year-RS for spermatocytic seminoma was close to 100% and the lowest 5-year-RS occurred among choriocarcinoma. Higher age at diagnosis is associated with a poorer prognosis among nonseminoma patients.     

Further studies of chimerism in heterosexual cattle twins

This study was designed to determine whether chimerism in heterosexual twin cattle could be detected in spleen and bone marrow and whether chimeric germ cells could survive into maturity and undergo meiosis. Differential erythrocyte typing and cytogenetic technics were employed. Somatic cell chimerism was usually equal in the various tissues examined. Germ cell chimerism was always low. Meiosis of XX germ cells in testis was detected through diakinesis. Lower ratios for germ cells than for somatic cells were obtained probably because they are mobile only for a short period of time in embryogeny, when they travel from yolk sac to gonadal primordia.     

Incidence patterns and trends of malignant gonadal and extragonadal germ cell tumors in Germany, 1998–2008

BackgroundMalignant gonadal (GGCT) and extragonal germ cell tumors [GCT (EGCT)] are thought to originate from primordial germ cells. In contrast to well reported population-based data of GGCTs in males, analyses of GGCTs in females and EGCTs in both sexes remain limited.MethodsIn a pooling project of nine population-based cancer registries in Germany for the years 1998–2008, 16,883 malignant GCTs and their topographical sites were identified using ICD-O morphology and topography for persons aged 15 years and older. We estimated age-specific and age-standardized incidence rates.ResultsAmong males, the incidence of testicular GCTs increased over time. In contrast, there was no increase in the incidence of EGCTs. Among females, rates of ovarian GCTs were stable, while rates of EGCTs declined over time. The most frequent extragonadal sites were mediastinum among males and placenta among females.ConclusionsOur results underline different incidence trends and distinct age-specific incidence patterns of malignant GGCTs and EGCTs, as reported recently by several population-based registries. The differences suggest that GGCT and EGCT may have different etiologies.     

miR-146a suppresses the sensitivity to interferon-α in hepatocellular carcinoma cells

Previous studies have demonstrated expression of Toll-like receptors (TLRs) in the surface epithelium of normal ovaries (OSE) and in epithelial ovarian tumors. Most notably, OSE-derived cancers express TLR4, which activates the nuclear factor-kappa B (NF-κB) signaling cascade as a mediator of inflammatory response. Currently, there is considerable interest in elucidating the role of TLR-mediated signaling in cancers. Nevertheless, the expression of TLRs in granulosa cell tumors (GCTs) of the ovary, and the extent to which GCT expression of TLRs may influence cell-signaling pathways and/or modulate the efficacy of chemotherapeutics, has yet to be determined. In the present study, human GCT lines (COV434 and KGN) were utilized to evaluate expression of functional TLR4. TLR4 is expressed in GCT cell lines and ligation of TLR4 with bacterial lipopolysaccharide (LPS) led to IκB degradation and activation of NF-κB. NF-κB activation was confirmed by nuclear localization of NF-κB p65 following treatment with LPS and the naturally occurring ligand, HSP60. Notably, immunoneutralization of TLR4 blocked nuclear localization, and inhibition of NF-κB signaling attenuated LPS-induced TNFα plus increased doubling time in both cell lines. Contradictory to reports using human OSE cell lines, inhibition of NF-κB signaling failed to sensitize GCT lines to TRAIL or cisplatin. In summary, findings herein are the first to demonstrate a functional TLR-signaling pathway specifically in GCTs, and indicate that in contrast to OSE-derived cancers, inhibition of NF-κB does not sensitize GCTs to TRAIL or cisplatin.     

Granular cell tumor of the biliary system: a report of 2 cases with cytologic diagnosis on endoscopic brushing

BACKGROUND: Granular cell tumors (GCTs) of biliary system are rare. GCTs show a striking preponderance for young, black females, who generally present with obstructive jaundice. To our knowledge, these are the first 2 reports of GCT of biliary system identifed on endoscopic brushing cytology. CASES: In case 1, a 24-year-old, black woman presented with a 5-month history of pruritus. Radiographic studies demonstrated a mass in the distal common bile duct. Endoscopic biopsy and bile duct brushing were diagnosed as GCT. A Whipple procedure was confirmatory of GCT. In case 2, a 38-year-old, black female presented with a 7-month history of pruritus and jaundice. Radiographic studies showed a stricture of the common hepatic duct at the hilum. Endoscopic brushing cytology of the stricture yielded only a few sheets of granular cells that were missed on initial screening. Suspicion of cholangiocarcinoma prompted surgery, and final histopathology showed GCT. Both patients were well 1 1/2 and 6 years after presentation. CONCLUSION: GCT of the bile duct can be diagnosed on endoscopic brushing and should be considered in the cytologic differential diagnosis in the appropriate clinical settings.