T cells in testicular germ cell tumors: new evidence of fundamental contributions by rare subsets.

BACKGROUND: Immune 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. METHODS: Human 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. RESULTS: Profound 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. CONCLUSION: Despite 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.

Hypomethylation-driven AKT Serine/Threonine Kinase 3 promotes testicular germ cell tumors proliferation and negatively correlates to immune infiltration.

AKT Serine/Threonine Kinase 3 (AKT3) has been reported to play an important role in different tumors. However, its clinical value, biological function, and molecular mechanism in testicular germ cell tumors (TGCT) remains unclear. In the current study, we applied the Gene Set Cancer Analysis (GSCA), UCSC XENA, Gene Expression Omnibus (GEO), the Human Protein Atlas (HPA), LinkedOmics, DiseaseMeth version 2.0, TISIDB, and other databases for TGCT data mining. Then, we investigated AKT3's mechanism of action and clinical survival significance via bioinformatics followed by in vitro experiments. We found that AKT3 was upregulated and had frequent copy number amplifications in TGCT, which were associated with poor survival outcomes of patients. On the other hand, mutations that led to AKT3 loss-of-function were correlated to a better prognosis in patients. Moreover, AKT3 silencing significantly inhibited the proliferation, DNA synthesis and colony formation of NCCIT cells (a TGCT cell line). AKT3 might participate in TGCT progression through multiple signaling pathways, such as ErbB, oxidative phosphorylation, and affecting tumor immune infiltration. Also, the upregulation of AKT3 mRNA expression might be driven by the hypomethylation of its promoter region. Overall, AKT3 is a potential TGCT oncogene and can be further used as a therapeutic target.

Are Changes in the Percentage of Specific Leukocyte Subpopulations Associated with Endogenous DNA Damage Levels in Testicular Cancer Patients?

Chemoresistance of germ cell tumors (GCTs) represents an intensively studied property of GCTs that is the result of a complicated multifactorial process. One of the driving factors in this process is the tumor microenvironment (TME). Intensive crosstalk between the DNA damage/DNA repair pathways and the TME has already been reported. This study aimed at evaluating the interplay between the immune TME and endogenous DNA damage levels in GCT patients. A cocultivation system consisting of peripheral blood mononuclear cells (PBMCs) from healthy donors and GCT cell lines was used in an in vitro study. The patient cohort included 74 chemotherapy-naïve GCT patients. Endogenous DNA damage levels were measured by comet assay. Immunophenotyping of leukocyte subpopulations was performed using flow cytometry. Statistical analysis included data assessing immunophenotypes, DNA damage levels and clinicopathological characteristics of enrolled patients. The DNA damage level in PBMCs cocultivated with cisplatin (CDDP)-resistant GCT cell lines was significantly higher than in PBMCs cocultivated with their sensitive counterparts. In GCT patients, endogenous DNA damage levels above the cutoff value were independently associated with increased percentages of natural killer cells, CD16-positive dendritic cells and regulatory T cells. The crosstalk between the endogenous DNA damage level and specific changes in the immune TME reflected in the blood of GCT patients was revealed. The obtained data contribute to a deeper understanding of ongoing interactions in the TME of GCTs.

Toll-like receptor 2 (TLR2) is a candidate prognostic factor in testicular germ cell tumors as well as an indicator of immune function in the tumor microenvironment.

Testicular cancer is the most common malignant tumor in young men, and its incidence has increased in recent years. The tumor microenvironment (TME) plays a crucial role in the development and progression of tumors; however, the TME of testicular germ cell tumor (TGCT) is poorly understood. In this study, we downloaded information for 156 TGCT cases from The Cancer Genome Atlas (TCGA) database, used the ESTIMATE method to determine immune and stromal scores, and used CIBERSORT to calculate the proportion of tumor-infiltrating immune cells (TICs). The differentially expressed genes were subjected to a COX regression analysis and used for the construction of a protein-protein interaction (PPI) network. Toll-like receptor 2 (TLR2) was identified as a predictive marker by combining the results of the Cox regression analysis and PPI network. A survival analysis showed that TLR2 was positively correlated with TGCT survival. A gene set enrichment analysis indicated that genes in the high TLR2 expression group were enriched for cell adhesion molecules (CAMs) and the chemokine signaling pathway, and genes in the low TLR2 expression group were mainly enriched in the spliceosome. Regarding proportions of TICs, naive B cells and follicular helper T cells were negatively correlated with the expression of TLR2. This suggests that as TLR2 expression increases, the immunocompetence of the TME decreases. The expression of TLR2 may affect the prognosis of TGCT, suggesting that this locus can be used as a prognostic factor for TGCT.

A Novel Artificially Humanized Anti-Cripto-1 Antibody Suppressing Cancer Cell Growth.

Cripto-1 is a member of the EGF-CFC/FRL1/Cryptic family and is involved in embryonic development and carcinogenesis. We designed a novel anti-Cripto-1 artificial antibody and assessed the recognition to the antigen and the potential to suppress the growth of cancer stem cells. First, single chain antibody clones were isolated by bio-panning with the affinity to recombinant Cripto-1 protein from our original phage-display library. Then, the variable regions of heavy chain VH and light chain VL in each clone were fused to constant regions of heavy chain CH and light chain CL regions respectively. These fused genes were expressed in ExpiCHO-S cells to produce artificial humanized antibodies against Cripto-1. After evaluation of the expression levels, one clone was selected and the anti-Cripto-1 antibody was produced and purified. The purified antibody showed affinity to recombinant Cripto-1 at 1.1 pmol and immunoreactivity to cancer tissues and cell lines. The antibody was available to detect the immunoreactivity in tissue microarrays of malignant tumors as well as in Cripto-1 overexpressing cells. Simultaneously, the antibody exhibited the potential to suppress the growth of human colon cancer derived GEO cells overexpressing Cripto-1 with IC50 at approximately 110 nM. The artificially humanized antibody is proposed to be a good candidate to target cancer cells overexpressing Cripto-1.

Leucine Zipper 4 Autoantibody: A Novel Germ Cell Tumor and Paraneoplastic Biomarker.

OBJECTIVE: This study was undertaken to describe a novel biomarker of germ cell tumor and associated paraneoplastic neurological syndrome (PNS). METHODS: Archival sera from patients with germ cell tumor-associated PNS were evaluated. We identified a common autoantigen in a human testicular cancer cell line (TCam-2) by Western blot and mass spectrometry. Its identity was confirmed by recombinant-protein Western blot, enzyme-linked immunosorbent assay (ELISA), and cell-based assay. Autoantibody specificity was confirmed by analyzing assorted control sera/cerebrospinal fluid. RESULTS: Leucine zipper 4 (LUZP4)-immunoglobulin G (IgG) was detected in 28 patients' sera, 26 of whom (93%) were men. The median age at neurological symptom onset was 45 years (range = 28-84). Median titer (ELISA) was 1:300 (1:50 to >1:6,400, normal value < 1:50). Coexistent kelchlike protein 11-IgG was identified in 18 cases (64%). The most common presenting phenotype was rhombencephalitis (17/28, 61%). Other presentations included limbic encephalitis (n = 5, 18%), seizures and/or encephalitis (n = 2, 7%), and motor neuronopathy/polyradiculopathy (n = 4, 14%). The most common malignancy among cancer-evaluated PNS patients was seminoma (21/27, 78%). Nine of the 21 seminomas detected by whole-body fluorodeoxyglucose positron emission tomography scan (43%) were extratesticular. Both female patients had ovarian teratoma. Regressed testicular germ cell tumors were found in 4 patients. Exposure of T-cell-dendritic-cell cocultures from chronic immunosuppression-naïve LUZP4-IgG-seropositive patients to recombinant LUZP4 protein evoked a marked increase in CD69 expression on both CD4+ and CD8+ T cells when compared to vehicle-exposed and healthy control cultures. INTERPRETATION: LUZP4-IgG represents a novel serological biomarker of PNS and has high predictive value for germ cell tumors. The demonstrated antigen-specific T-cell responses support a CD8+ T-cell-mediated cytotoxic paraneoplastic and antitumor potential. ANN NEUROL 2021;89:1001-1010.

Paraneoplastic Progressive Downbeat Nystagmus, Ataxia and Sensorineural Hearing Loss due to the ANTI-Kelch-11 Protein Antibody.

ABSTRACT: A 45-year-old man with a history of testicular seminoma treated 8 years earlier presented with chronic progressive truncal and limb ataxia, progressive sensorineural hearing loss, and episodic vertigo. Eye movement and neuro-otology examinations showed localizing abnormalities to the bilateral cerebellar flocculus, vermis, and bilateral cerebellar hemispheres. Audiometric testing showed bilateral symmetric sensorineural hearing loss. There was a normal MRI of the brain. Cerebrospinal fluid (CSF) showed modest lymphocytic pleocytosis, and there was an elevated serum choriogonadotrophic hormone. An abdominal CT scan showed a solitary, large retroperitoneal lymph node, and histopathologic examination of the node biopsy showed granulomatous inflammation without microorganisms; eventually, immunohistochemical markers confirmed the diagnosis of metastatic seminoma. Although normal neuroimaging and inflammatory CSF reaction suggested a paraneoplastic etiology, the initial paraneoplastic antibody testing was negative. Subsequent investigation identified a positive kelch-11 protein antibody, thus confirming the paraneoplastic connection between the metastatic seminoma and the subacute neurologic-cochleovestibular syndrome.

Promising Immunotherapy in Metastatic Testicular Sex Cord Stromal Tumours After First-Line Chemotherapy.

Background: Testicular sex cord stromal tumours (TSCSTs) are rare, with few studies focusing on the metastatic TSCST prognosis. The value of treatments, including radical orchiectomy (RO) and retroperitoneal lymph node dissection (RPLND), in preventing metastasis is controversial. Additionally, metastatic TSCSTs are resistant to chemotherapy. We aimed to assess the effectiveness and safety of immunotherapy in metastatic TSCSTs after first-line chemotherapy. Methods: We retrospectively screened patients with testicular tumours undergoing testis surgery between January 2005 and January 2019. Patients with TSCSTs who had undergone testis-sparing surgery (TSS) or RO were identified. The malignant type was defined as metastasis confirmed by pathology. Treatment responses, progression-free survival (PFS), overall survival (OS) and safety were analysed. Results: Among the 494 testicular tumour patients who received TSS or RO, 11 (2.2%) patients with histologically proven TSCSTs were identified. At the last follow-up, 7 patients survived without tumours, and 4 patients developed metastasis and received first-line cisplatin-based chemotherapy, with 1 of them achieving an objective response. Their PFS times were 1.5, 2.2, 9.0, and 17.0 months, respectively. Two patients received immune checkpoint inhibitors (ICIs) after developing chemotherapy resistance and achieved a partial response up to the last follow-up; one of them experienced Grade 1 adverse events, and the other experienced Grade 2 adverse events during immunotherapy. The median OS time of the 4 patients with metastatic TSCSTs was 32 months. Conclusions: TSCSTs are rare, and most are benign with a good prognosis. ICIs represent a promising option for improving clinical outcomes in metastatic TSCSTs.

Identification of novel genes in testicular cancer microenvironment based on ESTIMATE algorithm-derived immune scores.

Testicular cancer is the most common solid malignancy among young men. We downloaded data of testicular cancer patients from The Cancer Genome Atlas database to find novel genes in the testicular cancer microenviroment based on ESTIMATE algorithm-derived immune scores. A total of 156 cases of testicular cancer were included in this study and 165 cases of normal testicular tissues were used. We divided the testicular cancer patients into high- and low-score groups based on their immune scores. We identified 1,226 differentially expressed genes (fold change > 2, false discovery rate < 0.05), including 688 downregulated genes and 538 upregulated genes, between these two groups. The top Gene Ontology terms were involved in the immune response-regulating cell surface receptor signaling pathway, immune response-activating cell surface receptor signaling pathway, external side of the plasma membrane, and receptor ligand activity. By performing the Kyoto Encyclopedia of Genes and Genomes analysis, we demonstrated that cAMP signaling pathway was highly enriched among these differentially expressed genes. High expression of LINC01564, LINC02208, ODAM, RNA5SP111, and RNU6-196P were found to be associated with poor overall survival. The expression of genes was further validated by the Human Protein Atlas and only ALB and IFNG were demonstrated to be differentially expressed between testis tissue and testicular cancer tissue.

POTN: A Human Leukocyte Antigen-A2 Immunogenic Peptides Screening Model and Its Applications in Tumor Antigens Prediction.

Whole genome/exome sequencing data for tumors are now abundant, and many tumor antigens, especially mutant antigens (neoantigens), have been identified for cancer immunotherapy. However, only a small fraction of the peptides from these antigens induce cytotoxic T cell responses. Therefore, efficient methods to identify these antigenic peptides are crucial. The current models of major histocompatibility complex (MHC) binding and antigenic prediction are still inaccurate. In this study, 360 9-mer peptides with verified immunological activity were selected to construct a prediction of tumor neoantigen (POTN) model, an immunogenic prediction model specifically for the human leukocyte antigen-A2 allele. Based on the physicochemical properties of amino acids, such as the residue propensity, hydrophobicity, and organic solvent/water, we found that the predictive capability of POTN is superior to that of the prediction programs SYPEITHI, IEDB, and NetMHCpan 4.0. We used POTN to screen peptides for the cancer-testis antigen located on the X chromosome, and we identified several peptides that may trigger immunogenicity. We synthesized and measured the binding affinity and immunogenicity of these peptides and found that the accuracy of POTN is higher than that of NetMHCpan 4.0. Identifying the properties related to the T cell response or immunogenicity paves the way to understanding the MHC/peptide/T cell receptor complex. In conclusion, POTN is an efficient prediction model for screening high-affinity immunogenic peptides from tumor antigens, and thus provides useful information for developing cancer immunotherapy.

Immunosenescence profile and expression of the aging biomarker (p16INK4a) in testicular cancer survivors treated with chemotherapy.

BACKGROUND: Cytotoxic chemotherapy can cure advanced germ cell tumors. Nevertheless, cancer treatment may induce cellular senescence and accelerate molecular aging. The aging process implies an increase of cells expressing p16INK4a and changes in lymphocyte subpopulations. Our aim was to study the potential induction of premature immunosenescence in testicular cancer survivors (TCS) exposed to chemotherapy. METHODS: Case-control exploratory study of TCS treated with chemotherapy (≥3 BEP cycles, disease-free ≥3 months) compared with age matched healthy controls. Peripheral blood mononuclear cells were isolated, and lymphocyte subpopulations were analyzed by flow cytometry. CDKN2A/p16INK4a expression in T cells was measured using qPCR. The percentage of lymphocyte subpopulations and the CDKN2A/p16INK4a expression in TCS were compared with the control group using the Wilcoxon signed-rank test. RESULTS: We included 16 cases and 16 controls. The median age was 27 years (minimum 24, maximum 54) and the median time on surveillance was 26.5 months (minimum 3, maximum192). TCS had a lower percentage of total T cells and CD4+ T cells in total lymphocytes. Among the CD4+ T lymphocytes, TCS had less naïve CD4+ and increased memory CD4+ cells. Within the CD8+ T lymphocytes, TCS exhibited a decrease in the percentage of naïve cells and an increase in CD8 + CD45RA + CD57+ cells. TCS also exhibited decreased memory CD19+ B cells compared to the controls. The relative expression of CDKN2A/p16INK4a in T cells was increased in TCS (mean 1.54; 95% CI of the mean: 1.074-2.005; p = 0.048). CONCLUSION: In this exploratory study, TCS showed increased expression of CDKN2A/p16INK4a and a lymphocyte phenotype that has been associated with immunosenescence. Further studies are warranted to define the clinical implications of these alterations in TCS.

METTL3 May Regulate Testicular Germ Cell Tumors Through EMT and Immune Pathways.

Testicular germ cell tumors (TGCTs) are highly prevalent in young men aged 20-40 years and are one of the most common lethal solid tumors in men of this age. Due to the current unclear mechanism of tumor development, there is a lack of effective treatment, and therefore in-depth research of the molecular mechanism of the occurrence and development of TGCT and the search for suitable and effective therapeutic targets and molecular markers are of great significance for achieving effective treatment. METTL3 is a very important methylase, which has been implicated in the progression of many cancers, but the role of METTL3 in TGCT has not been fully elucidated. In this article, we found that METTL3 expression was significantly downregulated in TGCT tissues, and patients with low expression levels had lower overall survival and relapse-free survival rates. After overexpressing METTL3, cell proliferation, invasion, and migration ability significantly increased, while influencing the expression of epithelial-mesenchymal transition (EMT)-related proteins. In addition, we observed that the expression level of METTL3 positively correlated with molecular markers and infiltration level of CD8+ and CD4+ T cells and natural killer cells. In sum, our findings identified that METTL3 can be used as an independent prognostic marker in patients with TGCT. METTL3 participates in the proliferation, migration, and invasion of TGCT cells by regulating the expression of EMT-related genes and may also play a role in activating the tumor immune response in TGCT.

A model based on tumor-infiltrating immune cells for predicting the relapse rates of patients with testicular germ cell tumors.

OBJECTIVE: The activities of tumor-infiltrating immune cells (TIICs) play an important role in the outcomes of many types of cancers. Here, we sought to describe the landscape of TIICs in testicular germ cell tumors (TGCT) and to develop a prognostic model based on this information. METHODS: The Cell-type Identification by Estimating Relative Subsets of RNA Transcripts (CIBERSORT) algorithm was used to determine the proportions of 22 types of TIICs in a TGCT dataset (n = 74). Univariate and multivariate Cox regression analysis were used to develop an immune risk score (IRS) based on the association between TIICs and disease-free survival (DFS). The predictive accuracy of the IRS was evaluated using receiver operating characteristic curves, and the predictive accuracy of a prognostic nomogram was assessed using C-index and calibration curves. The biological functions of IRS-associated genes were evaluated by gene set enrichment analysis. RESULTS: The relative abundances of three TIICs (plasma cells, M2 macrophages, and resting mast cells) were significantly associated with DFS in TGCT patients. In receiver operating characteristic curve analysis, the resulting IRS had areas under the curve of 0.70, 0.793, and 0.827, for predicting 1-, 2-, and 3-year DFS, respectively. Kaplan-Meier analysis confirmed that DFS was shorter for patients with high IRS compared with low IRS. IRS was an independent predictor of disease recurrence (hazard ratio 1.306, 95% confidence interval 1.022-1.668; P = 0.033). The C-index for the nomogram was 0.733. Genes involved in cancer-associated and immunity-associated pathways were enriched in TGCT samples from patients in the high- and low-risk groups, respectively, and expression of four immune checkpoint regulators was significantly lower in the high IRS group compared with the low IRS group. CONCLUSIONS: A TIIC-based IRS may have utility as a complementary tool to predict relapse in patients with TGCT.

Cancer Stem Cell Niche and Immune-Active Tumor Microenvironment in Testicular Germ Cell Tumors.

Testicular germ cell tumors (TGCTs) represent the most common neoplasia among young men. Management of TGCTs is an excellent example of curative outcomes in clinical oncology. The unique sensitivity to cisplatin-based chemotherapy regimens has led to establishing TGCTs as a "model of cancer cure." However, mechanisms and factors underlying pervasive growth of TGCTs are still poorly understood. It is suggested that unique cancer stem cell (CSC) niche exists in the testicular tumor microenvironment. CSC niche potentially contributes to the progression of germ cell tumors. Furthermore, rich infiltration of TGCTs with immune cells indicates involvement of immune system in biology of this cancer type. This review summarizes current knowledge regarding specific cancer microenvironment in TGCTs and discusses the role of cancer stem cells as well as immune mechanisms in these tumors.

Lymphomas arising in immune-privileged sites: insights into biology, diagnosis, and pathogenesis.

Session 2 of the 2018 European Association of Hematopathology/Society for Hematopathology Workshop focused on lymphomas arising in immune-privileged sites: both lymphomas arising in the traditionally described "immune sanctuary" sites of the central nervous system (CNS) and testes, as well as those arising at sites of local immune privilege. Primary CNS large B cell lymphoma and primary testicular large B cell lymphoma were discussed, and the biology of these unique tumors was highlighted by several cases showing the classic mutation profile including MYD88 L265P and CD79B. The tendency of these tumors to involve both the CNS and testis was also reinforced by several cases. Four cases of low-grade B cell lymphomas (LGBCL) of the CNS were discussed. Two were classic Bing-Neel syndrome associated with LPL, and two were LGBCL with plasmacytic differentiation and amyloid deposition without systemic disease. Rare examples of systemic T and NK cell lymphomas involving the CNS were also discussed. Several cases of breast implant-associated anaplastic large cell lymphoma (BI-ALCL) were submitted showing the typical clinicopathologic features. These cases were discussed along with a case with analogous features arising in a patient with a gastric band implant, as well as large B cell lymphomas arising alongside foreign materials. Finally, large B cell lymphomas arising in effusions or localized sites of chronic inflammation (fibrin-associated diffuse large B cell lymphoma [DLBCL] and DLBCL associated with chronic inflammation) were described. The pathogenesis of all of these lymphomas is believed to be related to decreased immune surveillance, either innate to the physiology of the organ or acquired at a local site.

Malignant leydig cell tumor in a 91-year-old man: Case report.

Testicle tumors are a rare entity among men population, accounting for only 1-1.5% of all can-cers among men. The stromal tumors of the sexual cord correspond just 4% of all testicular cancers. Only 10% of them are malignant. The major representative of the sex cord-stromal tumors is the Leydig cell tumor, corresponding to 75 to 80% of the total. It has bimodal age incidence, involving children and adults between 30 and 60 years. We report the caso of a 91-year-old man with malignant Leydig cell tumor, presenting increase of the volume of scrotum, local pain and hyperemia. The are few cases in the literature, only 1 with pacient above 85 years old.

Malignant leydig cell tumor in a 91-year-old man: Case report

ABSTRACT Testicle tumors are a rare entity among men population, accounting for only 1-1.5% of all cancers among men. The stromal tumors of the sexual cord correspond just 4% of all testicular cancers. Only 10% of them are malignant. The major representative of the sex cord-stromal tumors is the Leydig cell tumor, corresponding to 75 to 80% of the total. It has bimodal age incidence, involving children and adults between 30 and 60 years. We report the caso of a 91-year-old man with malignant Leydig cell tumor, presenting increase of the volume of scrotum, local pain and hyperemia. The are few cases in the literature, only 1 with pacient above 85 years old.

May High Levels of Systemic Immune-Inflammation Index and Hematologic Inflammation Markers Suggest a Further Stage in Testicular Tumours?

INTRODUCTION: Due to variety of treatment alternatives for testicular tumours, parameters other than existing staging criteria are also needed. Most studies have revealed the correlation between cancer and inflammation. In this study, we aimed to investigate the value of preoperative inflammatory markers between early-stage testicular tumours and patients with advanced-stage, their relationship with tumour pathology and their importance in predicting stage. To calculate the differences between inflammatory markers, stage 1 tumours localized to the testis and advanced-stage tumours spread beyond the testis were classified into 2 groups according to tumour pathology. MATERIALS AND METHODS: The data of 112 patients undergoing inguinal orchiectomy in between 2008 and 2018 were recorded retrospectively. Neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), lymphocyte-to-monocyte ratio (LMR) and systemic immune-inflammation index (SII) were calculated by using the numbers of blood cell counts based systemic markers of inflammation. The differences between markers of inflammation were calculated by dividing tumours into 2 groups including early-stage and advanced- stage testicle tumours. RESULTS: According to the results of preoperative inflammatory markers in predicting the stage; in the seminoma group, the difference between the median NLR (2.37 vs. 4.39, p = 0.012), LMR (3.80 vs. 2.40, p = 0.018) and SII (612 vs. 1,127, p = 0.009) of stage 1 and advanced stage were statistically significant, while in the non-seminoma group, only the difference between median PLR (99 vs. 154, p = 0.002) of stage 1 and advanced stage was statistically significant. Sensitivity and specificity of predicting advanced stage according to cut-off values of markers were 69 and 75% in NLR (3.21), 83 and 75% in LMR, and 59 and 75% in SII in the seminoma group; on the other hand, in the non-seminoma group, the sensitivity, and specificity of predicting the advanced stage of PLR cut-off (104) were 71 and 88% respectively. CONCLUSIONS: The clinical use of inflammatory biomarkers in testicular tumours may represent an important step in understanding germ cell tumours biology and in supporting staging criteria and prognostic criteria.

High frequency of inactivating tetraspanin C D37 mutations in diffuse large B-cell lymphoma at immune-privileged sites.

Tetraspanin CD37 is predominantly expressed on the cell surface of mature B lymphocytes and is currently being studied as novel therapeutic target for B-cell lymphoma. Recently, we demonstrated that loss of CD37 induces spontaneous B-cell lymphoma in Cd37-knockout mice and correlates with inferior survival in patients with diffuse large B-cell lymphoma (DLBCL). Here, CD37 mutation analysis was performed in a cohort of 137 primary DLBCL samples, including 44 primary immune-privileged site-associated DLBCL (IP-DLBCL) samples originating in the testis or central nervous system. CD37 mutations were exclusively identified in IP-DLBCL cases (10/44, 23%) but absent in non-IP-DLBCL cases. The aberrations included 10 missense mutations, 1 deletion, and 3 splice-site CD37 mutations. Modeling and functional analysis of CD37 missense mutations revealed loss of function by impaired CD37 protein expression at the plasma membrane of human lymphoma B cells. This study provides novel insight into the molecular pathogenesis of IP-DLBCL and indicates that anti-CD37 therapies will be more beneficial for DLBCL patients without CD37 mutations.