The Testicular Cancer Consortium (TECAC): Filling Knowledge Gaps in the Genetic Etiology of Testicular Germ Cell Tumors.

Testicular seminoma and non-seminoma: ESMO-EURACAN Clinical Practice Guideline for diagnosis, treatment and follow-up

Testicular tumors are incredibly diverse and one of the most challenging areas in surgical pathology. Because of the rarity and overlapping features with numerous entities occurring in the testis and paratestis, these tumors pose a diagnostic challenge even to the most experienced general pathologists. In 2016, the latest "World Health Organization (WHO) classification of testicular tumors" was released, which incorporated several updates to the previous 2004 classification system. These updates involved several entities, including germ cell tumors, sex cord-stromal tumors, tumors containing both germ cells and sex-cord stromal cells, a miscellaneous group of testicular tumors and paratesticular tumors. In addition, significant changes were also introduced in the 2018 AJCC TNM staging (8th edition) regarding testicular tumors. The germ cell tumors are divided into 2 major groups; tumors derived from germ cell neoplasia in situ (GCNIS) and those unrelated to GCNIS. The GCNIS associated tumors include seminomatous and nonseminomatous germ cell tumors, which constitute a heterogeneous group of tumors. Non-GCNIS-associated tumors include prepubertal-type teratoma, prepubertal yolk sac tumor, mixed prepubertal-type teratoma and yolk sac tumor and spermatocytic seminoma. In the sex cord-stromal category, the tumors are classified based on their cells of origin. Most are Leydig cell tumors and Sertoli cell tumors; however, several mixed and diverse entities based on cell types are included in this group. Gonadoblastoma is the only tumor in the mixed germ cell and sex cord-stromal tumor category. Because of recent advances in molecular techniques, abundant new genetic information has emerged which helped classify the tumors based on the molecular alterations and provided insights into the tumor pathogenesis. This review focused on the updates related to testicular germ cell tumors and sex cord-stromal tumors and described the morphologic, immunohistochemical and molecular characteristics with an aim to provide a practical diagnostic approach and an update on relevant recent molecular advances.

The genomic landscape of testicular germ cell tumour (TGCT) can be summarized using four overarching hypotheses. Firstly, TGCT risk is dominated by inherited genetic factors, which determine nearly half of all disease risk and are highly polygenic in nature. Secondly KIT-KITLG signalling is currently the major pathway that is implicated in TGCT formation, both as a predisposition risk factor and a somatic driver event. Results from genome-wide association studies have also consistently suggested that other closely related pathways involved in male germ cell development and sex determination are associated with TGCT risk. Thirdly, the method of disease formation is unique, with tumours universally stemming from a noninvasive precursor lesion, probably of fetal origin, which lies dormant through childhood into adolescence and then eventually begins malignant growth in early adulthood. Formation of a 12p isochromosome, a hallmark of TGCT observed in nearly all tumours, is likely to be a key triggering event for malignant transformation. Finally, TGCT have been shown to have a distinctive somatic mutational profile, with a low rate of point mutations contrasted with frequent large-scale chromosomal gains. These four hypotheses by no means constitute a complete model that explains TGCT tumorigenesis, but advances in genomic technologies have enabled considerable progress in describing and understanding the disease. Further advancing our understanding of the genomic basis of TGCT offers a clear opportunity for clinical benefit in terms of preventing invasive cancer arising in young men, decreasing the burden of chemotherapy-related survivorship issues and reducing mortality in the minority of patients who have treatment-refractory disease.

SEXUAL FUNCTIONING AFTER MULTIMODALITY TREATMENT FOR DISSEMINATED NONSEMINOMATOUS TESTICULAR GERM CELL TUMOR

RISK ADAPTED MANAGEMENT WITH ADJUVANT CHEMOTHERAPY IN PATIENTS WITH HIGH RISK CLINICAL STAGE I NONSEMINOMATOUS GERM CELL TUMOR

Background: Testicular germ cell tumors (TGCT) are the most common cancers in young men of European ancestry aged 20 to 39 years. The incidence of TGCT has doubled over the past 20 years, yet no robust environmental risk factors for disease have been identified. Although TGCTs are treatable by surgery, radiation, and platinum-based chemotherapy, multiple long-term toxicities of treatment often occur impacting morbidity and mortality. Due to their high heritability and homogenous cell of origin, TGCTs are well suited to genome-wide association methods. The Testicular Cancer Consortium (TECAC) has brought together the largest genome-wise association study (GWAS) study of TGCT to date. Methods: We conducted a GWAS of 5,602 cases and 5,006 controls aggregated from 12 locations in the US and Europe. Logistic regression models adjusted for study center and genomic ancestry. Genotypes were imputed against the Human Haplotype Reference Consortium. Meta-analysis was performed to combine GWAS results with summary statistics from five previously published TGCT studies, UK Biobank, deCODE Genetics, and an independent set of cases and controls, for a total of 10,156 cases and 179,683 controls. Biologic function of loci was explored using PAINTOR annotated with ATAC-seq data of four TGCT cell lines, SPATIAL-seq data of the NTERA2 TGCT cell-line, and publicly available data from ENCODE. Polygenic risk scores (PRS) were computed using subject-level data from the 5,602 cases and 5,006 controls, and effect sizes of the novel hits derived from the meta-analysis. Results: 22 novel and 45 previously reported loci associated with TGCT surpassed genome-wide significance (p < 5e-08). We discovered additional markers in known susceptibility loci and identified novel regions associated with germ cell development and sex determination (e.g., BCL11, AR), immune function (e.g., TNXB, ITIH5), and for the first time identified genes associated with kinetochore activity (e.g., PPP2R5A, ANAPC2). All identified risk SNPs to date account for 42.3% of heritability. Men in the highest 1% of PRS had over a 15-fold increased risk of TGCT compared to those at the median PRS, and PRS overall had an AUC of 74.29%. Conclusions: Results from our TGCT meta-analysis continue to provide insights into biological pathways affecting germ cell specification, expression, and epigenetic reprogramming, and sex determination. Our results also uniquely place TGCT as the only cancer type in which inherited variants implicating kinetochore activity, critical for chromosomal segregation, have been identified. Citation Format: John Pluta, Louisa Pyle, Timothy Bishop, Javier Benitez, Victoria Cortessis, Alberto Ferlin, Jourik Gietema, Mark Greene, Thomas Grotmol, Ramneek Gupta, Robert Hamilton, Michelle Hildebrandt, Lambertus Kiemeney, Davor Lessel, Thorunn Rafnar, Lorenzo Richiardi, Rolf Skotheim, Clare Turnbull, Fredrik Wiklund, Tongzhang Zheng, Ewa Rajpert- De Meyts, Stephen Schwartz, Katherine McGlynn, Peter Kanetsky, Katherine Nathanson. Identification of 22 novel loci associated with susceptibility to testicular germ cell tumors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1203.

Testicular germ cell tumour (TGCT) is a malignancy with a high heritable component. The inherited risk is polygenic, and around 50 susceptibility genes are identified. The functional role of the gene products for TGCT development is not well understood. The focus of this review is functional studies of genetic risk factors for TGCT derived from GCNIS and the signalling pathways involved in the pathogenesis. Genome-wide association studies have identified new risk loci for TGCT and confirmed previously identified susceptibility genes. Many of these risk genes are related to male germ cell development, sex determination and genomic integrity. Gain- and loss-of-function studies in animal models and TGCT cell lines, as well as gene and protein expression studies in TGCT patient samples, have contributed to the understanding of TGCT development. KITLG-KIT signalling is of crucial importance, but several other signal transduction pathways may also play a role. Many of the risk loci are in non-coding regions, and studies have revealed that non-coding RNAs may act as oncogenes or tumour suppressors in TGCT development. The risk of TGCT is polygenic, and the underlying molecular mechanisms are complex. Several signalling pathways are related to TGCT development, and both proteins and non-coding RNAs may act as oncogenes or tumour suppressors. Epigenetic studies are of importance to get further knowledge about how the signalling pathways are regulated.

RETROPERITONEAL LYMPH NODE DISSECTION FOR THE MANAGEMENT OF CLINICAL STAGE I NONSEMINOMA

Integrating Metastasectomy in the Management of Advanced Urological Malignancies—Where are we in 2005?

Testicular germ cell tumour (TGCT) is the most common cancer in young men, and the incidence of TGCT is rising worldwide for unknown reasons [1], [2]. Treatments for TGCT are overall quite effective, but at the cost of significant toxicity [3], creating a powerful incentive for the development of more specific, molecularly guided therapies. TGCTs are generally thought to arise from a pluripotent fetal or embryonic germ cell [4]. Reflecting this pluripotency, these tumours can present in a wide range of histologic forms. Seminomas are TGCTs that retain features of pluripotent, primitive germ cells. In contrast, non-seminoma TGCTs exhibit differentiation into forms resembling somatic tissues (teratomas) or extraembryonic structures such as yolk sac (yolk sac tumour) or placenta (choriocarcinoma) (Figure 1). Family members of TGCT patients have a markedly increased risk of developing TGCT, strongly implicating an underlying genetic basis. Recent genome-wide association studies of TGCT have identified SNPs near ATF7IP, BAK1, DMRT1, KITLG, SPRY4, and TERT-CLPTM1L that increase TGCT risk [5]–[7]; however, the mechanisms associating most of these loci with tumourigenesis remain unclear. Figure 1 Animal models of TGCT and correlation to human histologic subtypes. Researchers seeking to identify such mechanisms have been hindered by the limited TGCT animal models available thus far. Two heritable TGCT models have been previously described: one in mouse and one in zebrafish. The mouse model arose through the observation by Leroy Stevens in the late 1950s that testicular teratomas arise spontaneously at low frequency during embryonic development in mice of the 129/Sv strain [8]. This discovery, which ultimately led to the experimental derivation of embryonic stem cells [9], [10], has also proved to be a useful model of teratoma formation. A number of genes have been identified as modifier loci that increase teratoma incidence in the 129/Sv background, including Tp53, Dmrt1, and Dnd1, an RNA-binding protein that is central to germ cell maintenance [11]–[13]. Forward genetic screening led to the discovery of a second in vivo TGCT model. Zebrafish carrying nonsense mutations in alk6b/bmpr1bb, an ortholog of the human bone morphogenetic protein (BMP) receptor BMPR1B, develop TGCTs resembling human seminomas [14], [15]. This finding illuminated the importance of BMP signaling in germ cell development and implicated disruption of BMP signaling in human germ cell tumourigenesis [16]. These two models have provided insight into the roles of pluripotency and differentiation pathways in TGCT development; however, their direct correlation to human tumourigenesis has been limited, as genes such as DND1 and BMPR1B have not been found to be mutated in human TGCTs [14], [17]. In this issue of PLOS Genetics, Basten and coworkers describe a new zebrafish TGCT model with a direct connection to human TGCT mutations [18]. Zebrafish with homozygous mutations in the ciliary protein lrrc50 were previously described to have kidney cysts homologous to human polycystic kidney disease [19]. In this paper, the authors report that male lrrc50 heterozygotes develop testicular tumours late in life with near complete penetrance. Morphologically, these tumours, similar to those arising in bmpr1bb-deficient zebrafish, contain sheets of uniform, undifferentiated germ cells, resembling human seminoma. Loss of heterozygosity at the lrrc50 locus was found in some tumours, consistent with a role of lrrc50 as a tumour suppressor. The authors then conducted a mutational analysis of LRRC50 in a collection of human seminoma samples and identified different mutations in two pedigrees with family history of seminomas, as well as heterozygosity for a different germline LRRC50 mutation in five of 38 patients with sporadic seminomas. LRRC50 is thus the first gene specifically linked to seminoma predisposition in humans. The mutations were found to be functional nulls through their inability to complement lrrc50 knockdown in zebrafish embryos, an elegant example of the utility of the fish system for both gene discovery (forward genetics) and functional genomics (reverse genetics). lrrc50 has heretofore been characterized solely as a ciliary motor protein, and its connection to GCT suppression is intriguing. Cilia have not previously been thought to be present in spermatogonia, but the authors show that normal spermatogonia do indeed have a cilium and that LRRC50 colocalized with the axoneme in spermatogonial stem cells. In addition, the authors provide evidence that its role may not be solely structural by showing that its expression is cell cycle–regulated and that it localizes with condensed chromosomes. The development of both renal cysts in homozygotes and seminomas in heterozygotes may implicate an underlying role for lrrc50 in early genitourinary development. Furthermore, the primary cilium has emerged as a signaling center for Hedgehog, Wnt, and other developmental pathways [20], and this, along with the TGCT phenotype of bmpr1bb mutants, raises the interesting possibility that interplay of these developmental signaling pathways is central to TGCT tumour suppression. Follow-up mechanistic studies will be critical to testing these hypotheses. This paper provides another example of the power of zebrafish forward genetic screens for discovery of genes with novel roles in cancer and other diseases, and is a welcome addition to the list of animal models of TGCT. Some caveats apply when comparing analogous tumours arising in animals separated by a large evolutionary distance; for example, the fish seminomas are benign compared to human seminomas, and may well arise at a different stage of germ cell development. More significantly, neither the fish nor the mouse models currently reflect the biology of human non-seminomatous GCTs, such as embryonal carcinoma, choriocarcinoma, or yolk sac tumour. Clinically the non-seminomas are more likely to be metastatic and resistant to standard treatments, meaning that new models of these GCT subtypes are urgently needed to provide insight into tumour biology, as well as platforms for testing new therapeutic strategies for these cancers.

The Y Deletion gr/gr and Susceptibility to Testicular Germ Cell Tumor

Multifocality in Testicular Germ Cell Tumors

Genome-wide association (GWA) studies of testicular germ cell tumor (TGCT) have identified 18 susceptibility loci, some containing genes encoding proteins important in male germ cell development. Deletions of one of these genes, DMRT1, lead to male-to-female sex reversal and are associated with development of gonadoblastoma. To further explore genetic association with TGCT, we undertook a pathway-based analysis of SNP marker associations in the Penn GWAs (349 TGCT cases and 919 controls). We analyzed a custom-built sex determination gene set consisting of 32 genes using three different methods of pathway-based analysis. The sex determination gene set ranked highly compared with canonical gene sets, and it was associated with TGCT (FDRG = 2.28 × 10(-5), FDRM = 0.014 and FDRI = 0.008 for Gene Set Analysis-SNP (GSA-SNP), Meta-Analysis Gene Set Enrichment of Variant Associations (MAGENTA) and Improved Gene Set Enrichment Analysis for Genome-wide Association Study (i-GSEA4GWAS) analysis, respectively). The association remained after removal of DMRT1 from the gene set (FDRG = 0.0002, FDRM = 0.055 and FDRI = 0.009). Using data from the NCI GWA scan (582 TGCT cases and 1056 controls) and UK scan (986 TGCT cases and 4946 controls), we replicated these findings (NCI: FDRG = 0.006, FDRM = 0.014, FDRI = 0.033, and UK: FDRG = 1.04 × 10(-6), FDRM = 0.016, FDRI = 0.025). After removal of DMRT1 from the gene set, the sex determination gene set remains associated with TGCT in the NCI (FDRG = 0.039, FDRM = 0.050 and FDRI = 0.055) and UK scans (FDRG = 3.00 × 10(-5), FDRM = 0.056 and FDRI = 0.044). With the exception of DMRT1, genes in the sex determination gene set have not previously been identified as TGCT susceptibility loci in these GWA scans, demonstrating the complementary nature of a pathway-based approach for genome-wide analysis of TGCT.

Does retroperitoneal lymphadenectomy for testicular germ cell tumor require a different approach in the presence of horseshoe kidney?

Contralateral biopsies in patients with testicular germ-cell tumour—nuisance or new sense?