Germinoma of medulla.

Germinoma occurring in the medulla oblongata is extremely rare. We report a case of primary intracranial germinoma arising in the medulla oblongata of a 24-year-old postpartum female who presented with progressive weakness of upper and lower limbs, seventh nerve palsy, and decreased palatal movements. Her MR imaging showed a heterointense mass lesion in the posterior portion of upper medulla, the histology of which was reported as germinoma. Germ cell tumors should be considered in the differential diagnosis of tumors occurring in the brain stem.

Optic nerve and chiasmal germinoma.

A 15-year-old boy presented with visual acuity of 20/200 OD and no light perception OS. The anterior segment of the left eye showed a relative afferent pupillary defect. A large (4.5 x 4.5 x 2.0 mm) infiltrative optic nerve head lesion with dilated vessels was seen OS with disc pallor OD. MRI of the brain and orbit revealed lobulated optic nerve thickening and chiasm. A biopsy revealed features consistent with germinoma and was positive for marker placental alkaline phosphatase. Systemic examination, chest x-ray, abdominal ultrasound, cerebrospinal fluid, and serology were normal. He received 27 Gy to the craniospinal region followed by a boost of 27 Gy to the left optic nerve. Eight months postirradiation, vision stabilization was achieved with 20/200 OD and light perception with inaccurate projection of rays OS.

Telomerase activity in germ cell cancers and mature teratomas.

BACKGROUND: An inverse relationship has been reported between the presence of telomerase enzymatic activity and the induction of differentiation in human tumor cell lines. Male germ cell tumors represent an attractive clinical model to assess this relationship further because high telomerase activity is present in normal germ cell progenitors and in embryonal carcinomas that can differentiate into mature teratomas. To investigate how telomerase activity and the differentiation state of germ cell tumors are related, telomerase activities and telomere lengths were measured in benign testicular tissues, germ cell cancers, and mature or immature teratomas. METHODS: By use of a modified telomeric repeat amplification protocol (TRAP) assay, telomerase activity was measured in four specimens of benign testicular tissue, in 27 germ cell cancers, in seven mature teratomas, and in one immature teratoma. Telomere lengths were measured in all specimens by restriction digestion of genomic DNA and Southern blot hybridization analysis. Associations between telomerase activity and tissue histopathology were assessed with two-sided Fisher's exact tests. RESULTS: Telomerase activity was detected in all examined germ cell cancers and in the benign testicular tissue specimens. In marked contrast, telomerase activity was not detected in any mature teratoma (P<.0001). Very long telomeres were detected in some mature teratomas, consistent with telomerase repression as a late event in teratoma formation. The immature teratoma, with malignant transformation, had high telomerase activity. CONCLUSION: Telomerase is active in germ cell cancers and repressed in mature teratomas. The absence of telomerase activity may contribute to the limited proliferative capacity of mature teratomas. These findings support the existence of an inverse relationship between telomerase activity and the differentiation state of clinical germ cell tumors.

Simultaneous detection of all four alkaline phosphatase isoenzymes in human germ cell tumors using reverse transcription-PCR.

We have developed a reverse transcription-PCR method that clearly distinguishes between the RNA transcripts of all four alkaline phosphatase (AP) genes. If compared to the methods used up to the present, the main advantages of the reverse transcription-PCR method presented are its specificity and high sensitivity. The germ cell AP and the placental AP, which are the two most closely related AP isoenzymes (98% homology), can clearly be distinguished without any interference by other AP isoenzymes. An enhanced expression of AP isoenzymes has been reported for various tumors. The examination of the pattern of AP isoenzyme expression in a specific tumor and the corresponding tissue of origin enables discrimination between eutopically and ectopically expressed isoenzymes and thus represents an important tool in the elucidation of AP isoenzymes as potential tumor markers. The pattern of AP expression in 15 germ cell tumors, 2 germinal epithelia adjacent to seminoma, 2 cell lines of germ cell tumor origin (Tera-1 and BeWo), and 5 normal testes was studied. In comparison to normal testes, in all seminomatous germ cell tumors eutopic expression of germ cell AP and ectopic expression of tissue-nonspecific AP were demonstrated. In both samples of pure embryonal carcinoma and in the embryonal carcinoma cell line, the transcription of all four mRNAs was shown. These results indicate that the expression of the isoenzymes depends on the degree of differentiation of a tumor and that a simultaneous up-regulation of all AP isoenzymes in all types of germ cell tumors does not exist.

Testisin, a new human serine proteinase expressed by premeiotic testicular germ cells and lost in testicular germ cell tumors.

We have cloned and characterized a cDNA encoding a new human serine proteinase, testisin, that is abundantly expressed only in the testis and is lost in testicular tumors. The testisin cDNA was identified by homology cloning using degenerate primers directed at conserved sequence motifs within the catalytic regions of serine proteinases. It is 1073 nucleotides long, including 942 nucleotides of open reading frame and a 113-nucleotide 3' untranslated sequence. Northern and dot blot analyses of RNA from a range of normal human tissues revealed a 1.4-kb mRNA species that was present only in testis, which was not detected in eight of eight testicular tumors. Testisin cDNA is predicted to encode a protein of 314 amino acids, which consists of a 19-amino acid (aa) signal peptide, a 22-aa proregion, and a 273-aa catalytic domain, including a unique 17-aa COOH-terminal hydrophobic extension that is predicted to function as a membrane anchor. The deduced amino acid sequence of testisin shows 44% identity to prostasin and contains features that are typical of serine proteinases with trypsin-like substrate specificity. Antipeptide antibodies directed against the testisin polypeptide detected an immunoreactive testisin protein of Mr 35,000-39,000 in cell lysates from COS-7 cells that were transiently transfected with testisin cDNA. Immunostaining of normal testicular tissue showed that testisin was expressed in the cytoplasm and on the plasma membrane of premeiotic germ cells. No staining was detected in eight of eight germ cell-derived testicular tumors. In addition, the testisin gene was localized by fluorescence in situ hybridization to the short arm of human chromosome 16 (16p13.3), a region that has been associated with allellic imbalance and loss of heterozygosity in sporadic testicular tumors. These findings demonstrate a new cell surface serine proteinase, loss of which may have a direct or indirect role in the progression of testicular tumors of germ cell origin.

Germ cell-like telomeric length homeostasis in nonseminomatous testicular germ cell tumors.

Telomere maintenance plays an important role in cell proliferation and tumor survival. Human male germ cells, which carry long telomeres and express telomerase, give rise to a highly heterogeneous group of malignant tumors. We compared telomeric length and telomerase activity between two major histological types of primary testicular germ cell tumors. Fifteen out of 16 seminoma samples revealed telomeric restriction fragment (TRF) length below 13 kb; the remaining seminoma showed a major TRF fraction of 18 kb and a distinct minor fraction of above 23 kb length. In contrast, all 13 samples from nonseminomas showed TRF length >/=23 kb, which is similar to that reported in human sperm. Nine out of 11 seminoma specimens and six out of seven nonseminomas studied showed moderate to high telomerase activity, the only telomerase-negative nonseminoma being pure mature teratoma. These results indicate to a major difference in telomeric length between seminomas and nonseminomas, which is apparently unrelated to the presence of telomerase activity, and suggest a germline-like homeostasis of telomeric length is preserved in human nonseminomas. Oncogene (2000) 19, 4075 - 4078.

Aneuploidy of human testicular germ cell tumors is associated with amplification of centrosomes.

Testicular germ cell tumors occur in three age groups. Seminomas and nonseminomas of adults, including mature teratomas, and the precursor carcinoma in situ (CIS) are aneuploid. This also holds true for yolk sac tumors of newborn and infants, while the mature teratomas of this age are diploid. In contrast, spermatocytic seminomas occurring in the elderly contain both diploid and polyploid cells. Aneuploidy has been associated with centrosome aberrations, sometimes related to overexpression of STK15. Aneuploidy of non-neoplastic germ cells has been demonstrated in the context of male infertility, a risk factor for the development of seminoma/nonseminoma. We investigated aneuploidy, centrosome aberrations and the role of STK15 in different types of testicular germ cell tumors as well as in normal and disturbed spermatogenesis. The aneuploid seminomas and nonseminomas tumors (including CIS) showed increased numbers of centrosomes, without STK15 amplification or overexpression. Four out of six infantile teratomas had normal centrosomes, the remaining two and an infantile yolk sac tumor showed a heterogeneous pattern of cells with normal or amplified centrosomes. Spermatocytic seminomas had two, four or eight centrosomes. Germ cells in seminiferous tubules with disturbed spermatogenesis shared both aneuploidy and centrosome abnormalities with seminomas/nonseminomas and showed a more intense STK15 staining than those with normal spermatogenesis and CIS. Therefore, aneuploidy of testicular germ cell tumors is associated with amplified centrosomes probably unrelated to STK15.

Resistance to platinum-containing chemotherapy in testicular germ cell tumors is associated with downregulation of the protein kinase SRPK1.

Male germ cell tumors (GCTs) are extremely sensitive to platinum-containing chemotherapy, with only 10% of patients showing therapy resistance. However, the biological basis of the high curability of disseminated GCTs by chemotherapy is still unknown. Recently, we demonstrated that the mammalian serine/arginine-rich protein-specific kinase 1 (SRPK1) is a cisplatin-sensitive gene, inactivation of which leads to cisplatin resistance. Because, in mammalians, the expression of SRPK1 is preferentially high in testicular tissues, cisplatin responsiveness of male GCTs might be associated with SRPK1 levels. In the present study, we monitored SRPK1 protein expression in a unique series of nonseminomatous GCTs by immunohistochemistry. Randomly selected GCTs (n = 70) and tumors from patients responding to standard chemotherapy (n = 20) generally showed strong SRPK1 staining. In contrast, expression in refractory GCTs (n = 20) as well as in GCTs from poor-prognosis patients responding to high-dose chemotherapy only (n = 11) was significantly lower (two-sided Wilcoxon rank sum test: P < .001). In conclusion, our data suggest that SRPK1 expression might be an important prognostic indicator for the chemoresponsiveness of nonseminomatous GCTs.

Aromatase expression in human germinomas with possible biological effects.

Gonadal aromatase expression has been demonstrated in human Leydig, granulosa, and thecal cells, but never in human germ cells. In an attempt to explain the unique occurrence of isosexual precocious puberty in a young girl with a hCG-secreting suprasellar germinoma, we demonstrated the presence of aromatase expression in the germ cell component of this tumor. Immunohistochemical staining for P450-aromatase and hCG using a peroxidase-labeled streptaviden-biotin technique was performed on tumor specimens from the above patient and from four other subjects with central nervous system germinoma. Cytoplasmic aromatase staining was present in the germinoma cells of four of five cases of central nervous system germinoma studied. Staining was absent in the lymphocytic element within the tumor and in negative control tissues. The demonstration of aromatase activity in the malignant element of human germinomas indicates that aromatase expression can occur in human germ cells after malignant transformation. This parallels the finding that the transformation of Sertoli cells to sex cord tumor with annular tubules in Peutz Jeghers syndrome is associated with the induction of marked aromatase expression and systemic estrogen effect. We propose that tumor aromatase played a similar role in the unique occurrence of isosexual precocity in a girl with a suprasellar germinoma.

Somatic isoform of angiotensin I-converting enzyme in the pathology of testicular germ cell tumors.

Retained fetal expression of angiotensin I-converting enzyme (ACE, CD143) has recently been shown in intratubular germ cell neoplasms (IGCN) and invasive germ cell tumors (GCT), suggesting the somatic isoform (sACE) as a characteristic component of neoplastic germ cells. We analyzed the distribution of sACE in 159 testicular GCT, including 87 IGCN. sACE protein was determined by immunohistochemistry (MAb CG2) on routinely formalin-fixed and paraffin-embedded tissue sections, supplemented by mRNA expression analysis using in situ hybridization. These data were compared with those obtained by germ cell/placental alkaline phosphatases (PIAP; MAbs PL8-F6 and 8A9) employing an uniform score system for the evaluation of immunoreactivity (IRS; possible values from 0 to 12). Expression of sACE and PIAP was found in all 87 analyzed IGCN (IRS > 4, median IRS of 12). Heterogeneous staining patterns were not related to the type of adjacent GCT but correlated with low expression in adjacent seminomas (P =.032 for sACE; P =.005 for PIAP). Both sACE and PIAP often showed a decreased and more heterogeneous but still moderate expression in 91 classic seminomas (median IRS of 8) and were completely absent in tumor cells of spermatocytic seminomas. Despite all similarities, we found sACE and PIAP differently regulated during GCT progression. This was documented by a well-preserved expression of either sACE or PIAP or both in all classic seminomas, low PIAP immunoreactivity in metastasis of seminomas, and completely diverging expression patterns in nonseminomatous GCT. Our findings underline the close molecular relationship between IGCN and seminoma, and suggest sACE as an appropriate marker for seminomatous differentiated tumors. HUM PATHOL 31:1466-1476.

Angiotensin I-converting enzyme and potential substrates in human testis and testicular tumours.

The angiotensin I-converting enzyme (ACE, kininase II, CD143) shows a broad specificity for various oligopeptides. Besides the well-known conversion of angiotensin I to II, ACE degrades efficiently kinins and the tetrapeptide AcSDKP (goralatide) and thus equally participates in the renin-angiotensin system, the kallikrein-kinin system, and the regulation of stem cell proliferation. In the mammalian testis, ACE occurs in two isoforms. The testicular isoform (tACE) is exclusively expressed during spermatogenesis and is generally thought to represent the germ cell-specific isozyme. However, we have previously demonstrated that, in addition to tACE, the somatic isoform (sACE) is also present in human germ cells. Similar to other oncofoetal markers, sACE exhibits a transient expression during foetal germ cell development and appears to be a constant feature of intratubular germ cell neoplasm, the so-called carcinoma-in-situ (CIS) and, in particular, of classic seminoma. This demands the existence of specific paracrine functions during male germ cell differentiation and development of male germ cell tumours, which are mediated by either of the two ACE isoforms. Considering the complexity of current data about ACE, a logical connection is required between (I) the precise localisation of ACE isoforms, (I) the local access to potential substrates and (II) functional data obtained by knockout mice models. The present article summarises the current knowledge about ACE and its potential substrates with special emphasis on the differentiation-restricted ACE expression during human spermatogenesis and prespermatogenesis, the latter being closely linked to the pathogenesis of human germ cell tumours.

A systematic review of lactate dehydrogenase isoenzyme 1 and germ cell tumors.

OBJECTIVES: To evaluate lactate dehydrogenase isoenzyme 1 (LD-1) as a tumor marker of germ cell tumors. METHODS: A literature search included a CancerLit and Medline computer search of articles regarding germ cell tumors and LD-1 published between 1963 to 99 and a manual search of reference lists, theses, and textbooks. Forty articles, letters to the editor, and abstracts on testicular germ cell tumors and 10 articles on ovarian germ cell tumors fulfilled inclusion criteria. RESULTS: Of 696 patients with testicular germ cell tumors, 423 (61%) had a raised serum LD-1 catalytic concentration (S-LD-1). Patients with seminoma have a raised S-LD-1 more often (63%) than those with nonseminoma (60%). S-LD-1 was raised less often in patients with stage I (48%) than in those with stage II (50%) and stage III (67%). S-LD-1, serum alpha fetoprotein concentration (S-AFP), and serum human chorionic gonadotropin concentration (S-hCG) were discordant. S-LD-1 predicted outcome in four studies: one study regarding relapse in patients with nonseminomatous testicular germ cell tumors stage I, and three studies regarding survival of patients with metastatic testicular germ cell tumors. In two of three studies, S-LD-1 was a better prognostic predictor for patients with metastatic testicular germ cell tumors than S-LD. Of 40 patients with ovarian germ cell tumors, thirty-five (88%) had a raised S-LD-1. CONCLUSIONS: S-LD-1 is a useful serum tumor marker of testicular germ cell tumors. For patients with ovarian germ cell tumors, S-LD-1 was raised more often than for patients with testicular germ cell tumors. Further studies are required for a general recommendation regarding the use of S-LD-1 for germ cell tumors.

A study of topoisomerase activity in human testicular cancers.

Topoisomerases are widely detected in rapidly proliferating cancer cells. Many anti-topoisomerase agents are utilized for cancer chemotherapy. Testicular cancers are highly chemotherapy sensitive, however, 10% of them are refractory to the standard regimen. To investigate the topoisomerase activities in human testicular neoplasms, we examined the activity of topoisomerase I (TopoI) and topoisomerase II (TopoII) in 29 testicular tumors. TopoI activity was observed irrespective of pathological types of the tumor (21/29). TopoII was detected in seminoma and teratocarcinoma (5/29). In our experience, seminoma was relatively sensitive to chemotherapy including anti-TopoII agents. Our results suggest that Topol inhibitors could be more effective against seminoma as well as the other types of testicular tumors.

Glutathione related enzymes in human testicular germ cell tumors and normal testes.

In this study, the activity of the glutathione related enzymes, namely glutathione S-transferase (GST), glutathione reductase (GSSG-R), Selenium-dependent and -independent glutathione peroxidase (GPX) of various TGC tumors (n = 18) obtained from untreated patients, was compared to that of the corresponding enzymes of normal testicular tissues (n = 5). The enzymes of all tumorous tissues except teratomas were significantly less active, than the corresponding enzymes of nontumorous tissues. The GST was in seminomas 4.3-20-, in embryonal carcinomas 47-, and in mixed tumors 13-47-fold less active than in the normal testes. The GST activity of teratomas was about half of that of the normal tissues. The Se-independent GPX, component of GST alfa class, comprised about 90 percent of the total GPX activity in normal testis; however it was absent or barely detectable in all TGC tumors except teratomas. The latter had about the same GPX activity as the tumor-free testicular tissues. Apart from the teratoma, the GSSG-R activity of all TGC tumors was also suppressed to about one third of that of the normal testis. The insufficient function of glutathione related enzymes of TGC tumors may contribute to their sensitivity against treatment. The poorer prognosis of teratomas, however, may be explained by the relatively higher activity of their detoxifying enzymes.

Differential expression of glutathione S-transferases in germ cell tumors of human testes.

Glutathione S-transferase (GST) isoenzymes alfa, mu and pi were assessed by Western blotting in normal testes, seminomas and non-seminomatous germ cell tumors (NSGCTs). GST alfa and mu were strongly expressed in all normal specimens (n = 6), the pi isoform, however, could not or barely be detected. Ten (92%) of 11 seminomas had GST pi and only 2 (16%) showed alfa expression. In contrast, twelve (80%) of 15 NSGCTs showed a significant level of GST alfa and only 2 (13%) had GST pi expression. The absence or paucity of mu isoform was characteristic for all neoplastic testicular tissues. A statistically significant relationship (P = 0.021) could be established between GST alfa and stages of disease, i.e., alfa isoform was prevalent in the later stage (IIB, III) NSGCT tumors. These results suggest that the poor prognosis of the later stage NSGCTs may be due to their high GST alfa content, while the GST pi does not seem to have role in this relation.

KIT (c-kit oncogene product) pathway is constitutively activated in human testicular germ cell tumors.

We investigated the expression of KIT (product of c-kit oncogene), gain-of-function mutations, and activation of its downstream signal transduction in human testicular cancers. KIT was expressed in 88% (22/25) of seminomas and in 44.4% (4/9) of non-seminomas compared to adjacent normal testicular tissue. Nine of the KIT-expressing seminomas had mutations (40.9%; 9/22) in the c-kit gene; two cases in exon 11 and 7 cases in exon 17. Two of these mutations in exon 17 were novel, and the other seven mutations were identical to the already known gain-of-function mutations which cause activation of KIT without ligand stem cell factor. All of the mutant KIT and 53.8% (7/13) of wild-type KIT were phosphorylated (activated) and associated with phosphorylated phosphatidylinositol 3-kinase (PI3K). Akt was also phosphorylated in these seminomas, suggesting that the KIT-PI3K-Akt pathway is activated in seminoma. These findings suggest that the KIT-PI3K-Akt pathway is constitutively activated in testicular germ cell tumors, due to overexpression of KIT protein and/or gain-of-function mutations in the c-kit gene.

Production of placental alkaline phosphatase (PLAP) and PLAP-like material by epithelial germ cell and non-germ cell tumours in vitro.

Placental and placental-like alkaline phosphatase (PLAP) levels in the culture media of 87 cell lines of neoplastic and 'normal' origin were measured by a conventional immunosorbent enzymatic assay (IAEA) and by a new immunoradiometric assay (IRMA). The IRMA detected immunoreactive PLAP in 37 of 80 (46%) human epithelial and germ cell cultures, while the IAEA detected PLAP in only 25 (33%). Of the 52 non-germ cell tumour cultures, the IRMA detected expression in 24 (46%) and the IAEA in only 16 (31%). In 17 cases (21%) the IRMA recorded levels double that of the IAEA, while in five cultures (6%) the reverse was true. The IRMA was much more robust than the IAEA and had considerably lower inter- and intra-assay coefficients of variation (3.75-8.5% vs 5.2-46%). Detection of PLAP(-like) expression by IAEA is dependent on neoplastic expression of enzymatically functional molecules and quantification assumes constant enzyme kinetics. PLAP-like material has a higher catalytic rate constant than PLAP and thus will give higher values on a stoichiometric basis in an IAEA. The higher detection rate and levels of PLAP-like material in neoplastic cultures when measured by the IRMA clearly demonstrate ectopic expression of non-enzymatic PLAP and PLAP-like genes. The incidence of PLAP(-like) expression by non-germ cell and possible germ cell tumours has been underestimated and its utility as a tumour marker should be re-examined using assays which measure antigen mass rather than phosphatase activity.

Loss of heterozygosity and decreased expression of NME genes correlate with teratomatous differentiation in human male germ cell tumors.

Embryonal carcinoma in the human male is a pluripotential germ cell tumor (GCT), which is suggested to further differentiate to teratoma which displays somatic differentiation representing all three germinal layers. In a panel of 37 GCTs we determined frequency of loss of heterozygosity (LOH) and the level of expression of nucleoside diphosphate kinase (NDPK) genes NME 1 and NME 2. The frequency of LOH in teratomas (86%) was found to be highly significant (P < 0.01) compared to embryonal carcinomas (17%). We also found that the NME encoded proteins are expressed at a 4-5 fold lower level in teratomas compared to embryonal carcinomas. These findings lead us to hypothesize that a critical level of NDPK may be necessary for suppression of aberrant somatic differentiation.

Tumor markers at the time of recurrence in patients with germ cell tumors.

BACKGROUND: alpha-fetoprotein (AFP), human chorionic gonadotropin (HCG), and lactate dehydrogenase (LDH) closely follow the course of germ cell tumors (GCTs) and are widely used for diagnosis, prognosis, and follow-up purposes. The objective of this study was to assess the concordance of tumor markers at the time of diagnosis and recurrence. METHODS: The authors reviewed the records of 794 patients with GCTs treated in three Spanish hospitals from 1977-1996 and analyzed the concordance between AFP, HCG, and LDH levels at diagnosis and first and second recurrence. A positive marker was defined as a level of AFP > 10 ng/mL, HCG > 5 IU/L, or LDH > the upper limit of normal. One hundred twenty-five patients were identified who developed a first recurrence (123 had marker levels recorded). The median age was 27 years (range, 14-78 years). Histology was seminoma in 36 patients (29%) and nonseminomatous GCT (NSGCT) in 87 patients (71%). RESULTS: Seventy-nine patients (64%) had elevated tumor markers at diagnosis and 76 (62%) at first recurrence. An elevated marker was present at first recurrence in 58 of 79 patients (73%) with initially positive markers and in 18 of 44 patients (41%) with initially negative markers. In 84 of 123 patients (68%), the same marker pattern (positive or negative) was present at the time of diagnosis and at first recurrence, 78% in seminomas and 64% in NSGCTs. The earliest indicator of recurrence was an elevated marker in patients with NSGCTs and a radiologic finding in patients with seminomas. Thirty patients developed a second recurrence, 27 of whom (90%) had the same marker pattern as at first recurrence. CONCLUSIONS: Tumor marker pattern at diagnosis is not a good predictor of the pattern at recurrence, particularly in patients with NSGCTs. Marker assessment should be included in the follow-up schedule regardless of levels at the time of diagnosis. Early detection of recurrence should not rely only on marker levels, even in patients with elevated levels at presentation.