Intratubular Germ Cell Neoplasia Research Articles

Testicular tumors in infants are rare, accounting for only 1-2% of pediatric solid tumors. Among these, teratomas are the most common type, particularly in the prepubertal age group. Unlike their post-pubertal counterparts, prepubertal-type mature teratomas are usually benign, lack malignant potential, and are remotely associated with germ cell neoplasia in situ (GCNIS). While mature teratomas are typically found in the abdomen, intratesticular prepubertal-type teratomas in infants are infrequent. The present study describes the case of an infant with an intratesticular mature teratoma. Five months old infant presented with painless right sided scrotal swelling detected incidentally. Scrotal ultrasound revealed a large anechoic cystic lesion measuring 28×18 mm with no septation, calcification, or solid component. Blood parameters including tumor markers viz. alpha-fetoprotein (AFP) and beta-human chorionic gonadotropin (β-hCG) were within normal range for age. Surgical tumor enucleation (testis sparing surgery) was performed, and the histopathological examination revealed a benign, prepubertal-type teratoma composed entirely of mature elements. Based on the findings diagnosis of prepubertal-type mature teratoma (MT) was thus made. Child was later was kept under observation in follow-up. Surgical intervention is commonly used for the management of benign testicular tumors in pediatric patients, including prepubertal teratomas. Indeed, testis-sparing surgery is a recommended option in prepubertal age patients with such a presentation of testicular mass with a benign ultrasonography finding like homogeneous, unilocular, non-septated cyst without any internal echoes, no calcification, no solid component and negative serum markers.

Germ Cell Neoplasia In Situ (GCNIS) is considered the precursor lesion for the majority of testicular germ cell tumors (TGCTs). The aim of this study was to evaluate whether first-order radiomics features derived from volumetric diffusion tensor imaging (DTI) metrics-specifically apparent diffusion coefficient (ADC) and fractional anisotropy (FA) histogram parameters-can detect GCNIS. This study included 15 men with TGCTs and 10 controls. All participants underwent scrotal MRI, including DTI. Volumetric ADC and FA histogram metrics were calculated for the following tissues: group 1, TGCT; group 2: testicular parenchyma adjacent to tumor, histologically positive for GCNIS; and group 3, normal testis. Non-parametric statistics were used to assess differences in ADC and FA histogram parameters among the three groups. Pearson's correlation analysis was followed by ordinal regression analysis to identify key predictive histogram parameters. Widespread distributional differences (p < 0.05) were observed for many ADC and FA variables, with both TGCTs and GCNIS showing significant divergence from normal testes. Among the ADC statistics, the 10th percentile and skewness (p = 0.042), range (p = 0.023), interquartile range (p = 0.021), total energy (p = 0.033), entropy and kurtosis (p = 0.027) proved the most significant predictors for tissue classification. FA_energy (p = 0.039) was the most significant fingerprint of the carcinogenesis among the FA metrics. These parameters correctly characterized 88.8% of TGCTs, 87.5% of GCNIS tissues and 100% of normal testes. Radiomics features derived from volumetric ADC and FA histograms have promising potential to differentiate TGCTs, GCNIS, and normal testicular tissue, aiding early detection and characterization of pre-cancerous lesions.

Several studies analyzed the "reprogramming" of germ cell tumors of the testis (GCTT), known to be an epigenetic process that results in the preservation of stem cell features and/or differentiation of GCTT. EZH2 is a methyltransferase involved in the epigenetic regulation of tumors and has become a promising therapeutic target, but few studies have analyzed its expression in GCTT, germ cell neoplasia in situ (GCNIS), and adjacent testis. We tested 131, 36, and 29 GCTT components, GCNIS, and adjacent testes, respectively. EZH2 expression was evaluated by H-score and compared between different subgroups by adopting median values and the Fisher exact test. We found that EZH2 was more highly expressed by adjacent testis/GCNIS rather than by GCTT (P < .001), with adjacent testis showing the highest values and being statistically significant compared to GCNIS (P < .001). In adjacent testis, EZH2 expression was mainly detected in spermatocytes (primary and secondary) and spermatids, with scattered positive spermatogonia. Seminoma/embryonal carcinoma showed statistically significantly higher EZH2 expression compared to the other nonseminomatous GCTT (P = .027). EZH2 is differentially expressed during GCTT reprogramming (adjacent testis [very high levels] → GCNIS [high levels] → seminoma/embryonal carcinoma [moderate levels] → other nonseminomatous GCTT [low/absent levels]), supporting its involvement in the epigenetic regulation for determining the fate of GCTT.

Pathologic and Oncologic Outcomes of Patients Undergoing Orchiectomy After Chemotherapy for Advanced Testicular Germ Cell Tumors.

Persistent Müllerian duct syndrome (PMDS) is a rare disorder of sex development in 46,XY individuals, characterized by the presence of Müllerian duct structures and typically associated with cryptorchidism and an elevated risk of testicular malignancy. Germ cell neoplasia in situ (GCNIS) is a known precursor to testicular germ cell tumors, but the earlier stage of pre-GCNIS remains poorly characterized, particularly in the context of PMDS. We report the first known case of pre-GCNIS identified in a patient with PMDS. A male infant presented with a non-palpable right testis and was found to have intra-abdominal gonads with Müllerian remnants. Histopathology revealed centrally located gonocytes with OCT3/4 expression but without the morphological features of GCNIS, consistent with pre-GCNIS. Management included right orchiectomy and left orchiopexy. This case highlights the underrecognized early stages of germ cell dysregulation in PMDS and raises important considerations about the timing of gonadectomy, fertility preservation, and long-term surveillance. Further research is needed to clarify the malignant potential of pre-GCNIS in DSD populations and to inform individualized, risk-based management strategies.

Spermatocytic tumor is a very rare clinical entity representing 1% of testicular tumors affecting elderly males with a men age of 52–59 years. It lacks features associated with tumors derived from germ-cell neoplasia in situ (GCNIS) including cryptorchidism and racial or ethnic predisposition. We present the case of an 89-year-old man with spermatocytic tumor admitted in the unit of urology with right-sided testicular swelling. In this paper we discuss the histopathology and ultrasound features of this testicular tumor.

Teratomas are broadly classified according to the presence or absence of germ cell neoplasia in situ (GCNIS), which is defined as neoplastic germ cell proliferation in the seminiferous tubules. Postpubertaltype teratoma is classified into a germ cell tumor derived from GCNIS, and it is malignant in adults. However, a prepubertal teratoma is classified as a germ cell tumor unrelated to GCNIS. It is common in children and has a benign course. We report a case in a 60-year-old man who had been aware of a nodule in his left scrotum for 15 years, and felt heaviness and pain from a few months before he visited a specialist. He visited a local urologist and was referred to our department under the diagnosis of a left testicular tumor in February 202X. In March 202X, he underwent a left high orchiectomy. A 12×5 mm white mass lesion was observed, and a pathological examination showed intestinal epithelial and bronchial epithelial structures. Additionally, no GCNIS was found in background seminiferous tubules. There was no amplification of chromosome 12 short arm as shown by the fluorescence in situ hybridization method. On the basis of these pathological findings, prepubertal-type teratoma was diagnosed in this patient.

Phenotypic and Molecular Features of Somatically Derived "Yolk Sac Tumors": Similarities and Differences With Counterparts of Germ Cell Origin.

Testicular microcalcifications consist of hydroxyapatite and have been associated with an increased risk of testicular germ cell tumors (TGCTs) but are also found in benign cases such as loss-of-function variants in the phosphate transporter SLC34A2. Here, we show that fibroblast growth factor 23 (FGF23), a regulator of phosphate homeostasis, is expressed in testicular germ cell neoplasia in situ (GCNIS), embryonal carcinoma (EC), and human embryonic stem cells. FGF23 is not glycosylated in TGCTs and therefore cleaved into a C-terminal fragment which competitively antagonizes full-length FGF23. Here, Fgf23 knockout mice presented with marked calcifications in the epididymis, spermatogenic arrest, and focally germ cells expressing the osteoblast marker Osteocalcin (gene name: Bglap, protein name). Moreover, the frequent testicular microcalcifications in mice with no functional androgen receptor and lack of circulating gonadotropins are associated with lower Slc34a2 and higher Bglap/Slc34a1 (protein name: NPT2a) expression compared with wild-type mice. In accordance, human testicular specimens with microcalcifications also have lower SLC34A2 and a subpopulation of germ cells express phosphate transporter NPT2a, Osteocalcin, and RUNX2 highlighting aberrant local phosphate handling and expression of bone-specific proteins. Mineral disturbance in vitro using calcium or phosphate treatment induced deposition of calcium phosphate in a spermatogonial cell line and this effect was fully rescued by the mineralization inhibitor pyrophosphate. In conclusion, testicular microcalcifications arise secondary to local alterations in mineral homeostasis, which in combination with impaired Sertoli cell function and reduced levels of mineralization inhibitors due to high alkaline phosphatase activity in GCNIS and TGCTs facilitate osteogenic-like differentiation of testicular cells and deposition of hydroxyapatite.

Testicular microcalcifications consist of hydroxyapatite and have been associated with an increased risk of testicular germ cell tumors (TGCTs) but are also found in benign cases such as loss-of-function variants in the phosphate transporter SLC34A2. Here, we show that fibroblast growth factor 23 (FGF23), a regulator of phosphate homeostasis, is expressed in testicular germ cell neoplasia in situ (GCNIS), embryonal carcinoma (EC), and human embryonic stem cells. FGF23 is not glycosylated in TGCTs and therefore cleaved into a C-terminal fragment which competitively antagonizes full-length FGF23. Here, Fgf23 knockout mice presented with marked calcifications in the epididymis, spermatogenic arrest, and focally germ cells expressing the osteoblast marker Osteocalcin (gene name: Bglap, protein name). Moreover, the frequent testicular microcalcifications in mice with no functional androgen receptor and lack of circulating gonadotropins are associated with lower Slc34a2 and higher Bglap/Slc34a1 (protein name: NPT2a) expression compared with wild-type mice. In accordance, human testicular specimens with microcalcifications also have lower SLC34A2 and a subpopulation of germ cells express phosphate transporter NPT2a, Osteocalcin, and RUNX2 highlighting aberrant local phosphate handling and expression of bone-specific proteins. Mineral disturbance in vitro using calcium or phosphate treatment induced deposition of calcium phosphate in a spermatogonial cell line and this effect was fully rescued by the mineralization inhibitor pyrophosphate. In conclusion, testicular microcalcifications arise secondary to local alterations in mineral homeostasis, which in combination with impaired Sertoli cell function and reduced levels of mineralization inhibitors due to high alkaline phosphatase activity in GCNIS and TGCTs facilitate osteogenic-like differentiation of testicular cells and deposition of hydroxyapatite.

Untreated cryptorchid testes are in risk of intratubular germ cell neoplasia and subsequently may give rise to invasive germ cell tumors. Tissue samples were obtained from patients undergoing orchidectomy or orchiopexy and were subjected to routine histopathological and immunohistochemical examinations. Forty-three patients were enrolled in this study out of which 30 samples were collected. The mean age of patients was 9.16 years. One case (6.7%) showed positivity of the germ cell neoplasia with anto placental alkaline phosphatase and Anto CD-117 positive. Other histopathological findings such as fibrocollagenous tissue and Leydig cell hyperplasia were reported. Early surgical management is of importance for a better outcome in cases of undescended testes.

Abstract K. Bangalore Krishna: None. F.X. Schneck: None. M. Reyes-Mugica: None. S.A. Yatsenko: None. S.F. Witchel: None. Introduction: Individuals with 45,X/46,XY karyotypes have increased risks of developing germ cell tumors such as gonadoblastoma, defined as an in situ form of malignant germ cell tumor consisting of germ cell neoplasia in situ (GCNIS) / seminoma / dysgerminoma cells and incompletely differentiated sex cord cells reminiscent of Sertoli/granulosa cells. Germ cell tumor prevalence is 15-40% in patients with 45,X/46,XY mosaicism. We have previously reported bilateral gonadoblastoma in a 3 month old 45,X/46,XY child (PMID: 30646821). Dissecting gonadoblastoma of the ovary is characterized by a unique distribution of neoplastic germ cells. We report a child with bilateral dissecting gonadoblastoma. Clinical Case: This full-term neonate featured a 11x8 mm phallic structure, mildly rugated and fused labioscrotal folds, and a palpable gonad in the right inguinal canal. FISH analysis of 510 cells from a peripheral blood sample showed 507 cells with monosomy X and 3 cells with X and Y sex chromosomes. FISH studies on urine cells showed concordant results. Ultrasound revealed normal kidneys, an atypical appearing uterus, and bilateral oval hypoechoic to isoechoic structures lacking definite follicles in both adnexa. Shared decision-making discussions with parents concluded with decision for female sex of rearing. At 8 weeks of age, the phallus had lengthened and testosterone level was 120 ng/dl. Due to risk for germ cell tumors, bilateral gonadectomy was performed at 8 months of age. No further phallic growth was noted. Pathology showed left streak gonad with undifferentiated gonadal tissue/dissecting gonadoblastoma; right gonad with streak testis with undifferentiated gonadal tissue/dissecting gonadoblastoma. Fallopian tubes and associated Mullerian/Wolffian derivatives were identified. Cytogenetic analysis showed two cell lines: 45,X and another cell line with a structurally abnormal Y chromosome containing a duplication of the Yq11.23-qter segment. Staining showed numerous SALL4+ cells, some were also positive for OCT4, PLAP, C-Kitt and PgP9.5 stains. Conclusions: This infant’s phenotype and karyotype were troubling due to her risk for germ cell neoplasia. Based on our prior finding of bilateral gonadoblastoma in a 3-month infant, gonadectomy was performed at 8 months of age. Classical gonadoblastoma is considered to be non-invasive requiring only gonadectomy as treatment. Dissecting gonadoblastoma describes a pattern with either an infiltrative type or diffuse pattern instead of the typical small, nested arrangement. Dissecting gonadoblastoma can be confused with germinoma (PMID: 27454939). Correct identification of the histology is essential to avoid the more aggressive treatments often needed for germinoma. Monday, June 3, 2024

(273) COMPARATIVE RATES OF INCIDENTAL GERM CELL NEOPLASIA <i>IN SITU</i> IN HISPANIC MEN PRESENTING FOR INFERTILITY

BackgroundTesticular germ cell tumor (TGCT) is the most common type of tumor in young men. Type II germ cell tumors including postpubertal‐type teratomas are derived from the germ cell neoplasia in situ (GCNIS), whereas prepubertal‐type teratomas arise independently of the GCNIS. The consomic mouse strain 129.MOLF‐Chr19 (M19) is a suitable murine model of such tumors, but its characterization remains incomplete.ObjectiveHere, we interrogated the suitability of testicular tumors in M19 mice as a model of human TGCT by analyzing their histological features and gene expression signature.Material and methodsTestes collected from M19 mice of different ages were categorized by macroscopic appearance based on size and the degree of suspected tumorigenesis. Histological sections from selected tumors were stained with Hematoxylin and Eosin, and expression of genes associated with tumorigenesis was determined in frozen tissue samples from a large range of tumors of different subclasses using RT‐qPCR and Fluidigm Dynamic Arrays.ResultsMacroscopically, testicular specimens appeared very heterogeneous concerning size and signs indicating the presence of a tumor. Histological analysis confirmed the development of teratomas with areas of cells corresponding to all three germ cell layers. Gene expression analyses indicated upregulation of markers related to proliferation, vascular invasive potential and pluripotency, and revealed a strong correlation of gene expression with tumor size and a significant intercorrelation of individual genes.Discussion and conclusionTGCT in M19 mice is reminiscent of human testicular teratomas presenting with areas of cells derived from all germ layers and showing a typical gene signature. We thus confirm that these mice can serve as a suitable murine model of pure teratomas for preclinical research.

Background and Objectives: The presence and contribution of senescent cells in premalignant lesions is well documented, but not in germ cell neoplasia in situ. The purpose of this study is to identify the presence of senescent cells in pre-malignant testicular conditions and in different histological types of testicular cancer. Materials and Methods: Thirty patients who underwent orchiectomy due to testicular tumors were included. Formalin-fixed paraffin-embedded (FFPE) testicular tissue for each patient was available. Sections from these specimens were examined by immunohistochemical analysis with the following markers: GL13 for cellular senescence, p21WAF1/Cip1 for cell cycle arrest, and Ki67 for cell proliferation. Results: Thirteen (43.3%) suffered from seminoma with a mean total proportion of GCNIS senescence of 20.81 ± 6.81%. In the group of embryonal testicular tumors, nine (30%) patients were included, with an average rate of 6.64 ± 5.42% of senescent cells in GCNIS. One (3.3%) patient suffered from chondrosarcoma in which 7.9% of GL13+ cells were detected in GCNIS. Four (13.4%) patients suffered from teratoma and three (10%) from yolk sac tumors, while GCNIS senescence was detected in a range of 4.43 ± 1.78% and 3.76 ± 1.37%, respectively. Conclusions: Cellular senescence was detected in both germ cell neoplasia in situ and testicular cancer, but was more prevalent within the premalignant lesions.

Testicular germ cell tumours (GCT) are divided into three different subtypes (types I-III) regarding to their developmental origin, histological differences and molecular features. Type I GCT develop from disturbed primordial germ cells and most commonly occur in children and young adolescents, which is why they are referred to as prepubertal GCT. Type II GCT develop from a non-invasive germ cell neoplasia in situ (GCNIS) and show an isochromosome 12p (i12p) or gain of 12p material as a common and characteristic molecular alteration. Type III GCT originate from distorted postpubertal germ cells (e.g. spermatogonia) in adult patients and have changes on chromosome 9 with amplification of the DMRT1 gene. Type I GCT encompass prepubertal-type teratomas and yolk-sac tumours (YST). Type II GCT include seminoma, embryonal carcinoma, choriocarcinoma, postpubertal-type teratoma and postpubertal-type YST. Types I and II GCT both show similar morphology, but are separated from each other by the detection of a GCNIS and an i12p in type II GCT. For type II GCT it is especially important to detect non-seminomatous elements, as these tumours have a worse biological behaviour and need a different treatment to seminomas. In contrast to types I and II GCT, type III tumours are equivalent to spermatocytic tumours and usually occur in elderly men, with few exceptions in young adults. The development of types I and II GCT seems to depend not upon driver mutations, but rather on changes in the epigenetic landscape. Furthermore, different pluripotency associated factors (e.g. OCT3/4, SOX2, SOX17) play a crucial role in GCT development and can be used as immunohistochemical markers allowing to distinguish the different subtypes from each other in morphologically challenging tissue specimens. Especially in metastatic sites, a morphological and immunohistochemical diagnostic algorithm is important to detect small subpopulations of each non-seminomatous GCT subtype, which are associated with a poorer prognosis and need a different treatment. Furthermore, primary extragonadal GCT of the retroperitoneum or mediastinum develop from misguided germ cells during embryonic development, and might be challenging to detect in small tissue biopsies due to their rarity at corresponding sites. This review article summarises the pathobiological and developmental aspects of the three different types of testicular GCT that can be helpful in the histopathological examination of tumour specimens by pathologists.

Beckwith-Wiedemann spectrum (BWSp) is caused by genetic and epigenetic alterations on chromosome 11 that regulate cell growth and division. Considering the diverse phenotypic landscape in BWSp, the characterization of the CDKN1C molecular subtype remains relatively limited. Here, we investigate the role of CDKN1C in the broader BWSp phenotype. Notably, patients with CDKN1C variants appear to exhibit a different tumor risk than other BWSp molecular subtypes. We performed a comprehensive literature review using the search term "CDKN1C Beckwith" to identify 113 cases of patients with molecularly confirmed CDKN1C-BWSp. We then assessed the genotype and phenotype in a novel cohort of patients with CDKN1C-BWSp enrolled in the BWS Research Registry. Cardinal and suggestive features were evaluated for all patients reported, and tumor risk was established based on available reports. The most common phenotypes included macroglossia, omphalocele, and ear creases/pits. Tumor types reported from the literature included neuroblastoma, acute lymphocytic leukemia, superficial spreading melanoma, and intratubular germ cell neoplasia. Overall, this study identifies unique features associated with CDKN1C variants in BWSp, enabling more accurate clinical management. The absence of Wilms tumor and hepatoblastoma suggests that screening for these tumors may not be necessary, while the neuroblastoma risk warrants appropriate screening recommendations.

Embryonic-type neuroectodermal elements are often intimately mixed with primitive endodermal-type glands, like those of yolk sac tumors, in germ cell neoplasia in situ (GCNIS)-derived germ cell tumors of the testis. Because the primitive glands mimic tubules or rosettes of embryonic-type neuroectodermal elements, these embryonic-type neuroectodermal/glandular complexes may be misinterpreted as pure lesions of embryonic-type neuroectodermal elements, which, if of sufficient size, may lead to a diagnosis of embryonic-type neuroectodermal tumor, despite that the criteria of the World Health Organization for a "somatic-type malignancy" are not met. A diagnosis of embryonic-type neuroectodermal tumor in the testis may lead to retroperitoneal lymphadenectomy even in clinical stage I patients, and in postchemotherapy resections indicates a poor prognosis. The distinction of the neuroectodermal and glandular elements is not always straightforward based on morphology alone. We, therefore, studied 34 testis-derived germ cell tumors with embryonic-type neuroectodermal/glandular complexes and 2 purely glandular yolk sac tumors to characterize the immunophenotypes and determine an efficient immunohistochemical panel to aid in this differential. We found that GFAP, synaptophysin, and paired-like homeobox 2B (PHOX2B) expression was specific to embryonic-type neuroectodermal elements, although PHOX2B had poor sensitivity. In contrast, positive reactions with antibodies directed against AFP, villin, and CDX2 were specific for the glandular elements, although CDX2 had poor sensitivity. Other markers, including AE1/AE3 cytokeratin, SALL4, glypican 3, SOX2, SOX11, CD56, INSM1, and neurofilament, proved less helpful because of their nonspecificity and/or poor sensitivity. We conclude that the optimal immunohistochemical panel for distinguishing the components of embryonic-type neuroectodermal/glandular complexes includes stains for synaptophysin, GFAP, villin, and AFP.

BackgroundImmune cell infiltration is heterogeneous but common in testicular germ cell tumors (TGCT) and pre-invasive germ cell neoplasia in situ (GCNIS). Tumor-infiltrating T cells including regulatory T (Treg) and follicular helper T (Tfh) cells are found in other cancer entities, but their contributions to TGCT are unknown.MethodsHuman testis specimens from independent patient cohorts were analyzed using immunohistochemistry, flow cytometry and single-cell RNA sequencing (scRNA-seq) with special emphasis on delineating T cell subtypes.ResultsProfound changes in immune cell composition within TGCT, shifting from macrophages in normal testes to T cells plus B and dendritic cells in TGCT, were documented. In most samples (96%), the CD4+ T cell frequency exceeded that of CD8+ cells, with decreasing numbers from central to peripheral tumor areas, and to tumor-free, contralateral testes. T cells including Treg and Tfh were most abundant in seminoma compared to mixed tumors and embryonal carcinoma.ConclusionDespite considerable heterogeneity between patients, T cell subtypes form a key part of the TGCT microenvironment. The novel finding of rare Treg and Tfh cells in human testis suggests their involvement in TGCT pathobiology, with implications for understanding tumor progression, to assess patients’ prognosis, and as putative targets for personalized immunotherapy.

Cryptorchidism in males constitutes a notable risk factor for both infertility and testicular cancer. Infertility in adulthood is closely linked to the germ cell status in childhood. Furthermore, the significance of germ cell status is important as more than 95% of all reported testicular malignancies are germ cell tumors. The review aims to elucidate the pathogenesis of germ cells in cryptorchid testes concerning their association with infertility and testicular malignancies. Impaired germ cell numbers are evident in cryptorchid testes even during antenatal and neonatal stages. In cryptorchidism there is a rapid decline in germ cell number within the first year of life, partially attributed to physiologic gonocyte apoptosis. Additionally, germ cells fail to differentiate normally during mini-puberty leading to reduced germ cell proliferation and delayed clearance of gonocytes from the seminiferous epithelium. Absence of germ cells in testicular biopsies occurs already 10 months of age and germ cell deterioration progressively worsens with approximately 50% of persisting cryptorchid testes lacking germ cells during puberty. The deficient germ cell maturation and proliferation leads to later infertility. Elevated temperature in the cryptorchid testes and also hormonal deficiency contribute to this phenomenon. Germ cell neoplasia in situ (GCNIS) originating during fetal development may manifest in rare cases associated with disorders of sexual development, chromosomal abnormalities in boys, specific syndromes, and teratomas that include cryptorchidism. In adults, the presence of GCNIS predominantly represents a new histology pattern before invasive germ cell cancer is demonstrated and is neither congenital nor related to abnormal gonocyte transformation.